John Hopkins University v. 454 Life Sciences Corporation

Filing 119

MEMORANDUM OPINION re bench trial decision. Signed by Judge Leonard P. Stark on 1/26/17. (ntl)

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IN 'THE 'UNITED STATES DISTRICT 'COURT FOR 5fUEDISTRICfOF DELAWARE JOHNS HOPKINS UNNERSHY, Plaintiff, v. C.A.No. 13-:-1'853-LPS . 454 LIFE SCIENCES'lCORPORATION, Defendant. · KarenL Pascale,JamesL.'Higgins, YOUNG·CONAWAYSTARGATT& TAYLORLLP, Wilmington, DE RobertF. Altherr,Jr.,Joseph M. Skerpon, Christopher B.~Roth, BANNER& WITCOFF, LTD., .Was~ington, DC .Attorneys Jor .PlaintiffJohns Hopkins University. Kelly E. Farnan, Selena E. 'Molina, RICHARDS, .LAYTON & FINGER, P.A., Wilmington, DE R. Danny Huntington, Sharon E. Crane, C. Nichol~ Gifford, :Seth E. ·Cockrum, ROTHWELL, 'FIGG, ERNST.& MANBECK P.C., Washington, DC ·Attorneys for Defendant 454 .Life ·Sciences ·Corporation. MEMORANDUM OPINION : I January 26, 2017 Wilmington, Delaware On November 6, 2013., Johns Hopkins University ("JHU" or "Plaintiff') filed this :action pursuant to 35 U.S.C. § 146, seeking review of the DeCision and Final Judgmentofthe Board of Patent Appeals and Interferences ("the Board") in Interference No. 105,857 ("the Interference.,') .. · (See D.I. 1 at l, 8; Statement of Admitted Facts ("SAF") D.I. 102-1 ··if 1) The interfering applications are JHU's U.S. Patent Application No. 12/361,690 (D.I . .39 Ex. 1) (JHU's "'690 application") and U.S. Patent Application No. 13/33,240 (D.I. 39 Ex. 5) (454's "'240 application"), which was filed by Defendant 454 Life Sciences Corporation ('·'454" or "Defendant"). The Illterference involves a single .count ("Count"),1 with the interfering subject matter represented by claim 1 ·ofJHU's '690 application and claim 52 of 454's '240 application. (See D.I. 44 at l; D.I. 45at1) ·Claim 1 ofJHU' s '690 application recites the following four-step method:. A method for analyzing nucleic acid sequel}ces comprising: (a} generating a plurality_of molecules of a fragment of deoxyribonucleic acid; (b) delivering the plurality of molecules of the fragment of deoxyribonueleic _acid into aqueous. microreactors in a water-in-oil emulsion such that a plurality of aqueous microreactors comprise a single molecule of the fragment of deoxyribonucleic acid, ·a single· bead capable of hybridizing the fragment of deoxyribonucleic acid, and reagents necessaryto perform deo~yribonucleic acid amplification; 1 In an interference, "[t]he applicant mustidentify at least one patentable claim from every application or patent that interferes for each count. A count is just a description of the interfering subject matter, which the Board of Patent Appeals and Interferences uses to determine what evidence may be used to prove priority under 35 U.S.C. § 102(g)(l)." Manual of Patent Examining Procedure ("MPEP") § 2304.02(b). . 1 · (c) amplifying the fragment of deoxyribonucleic . acid in the microreactors to form amplified .copies of said fragment .ofdeoxyribonudeic acid bound':to beads in the niicroreactors; '[and] ( d].detetmining presence of amplified copies of said fragment. of deo~yribonudeic acid ·bound to a bead. . ·(DJ. 44 at 2-3)2 The Courtbdd a claim constructionhearing onJune 9,2015~andissued.amemorandum opiriion on.daim,construction on August 24, 2015. (D.I. 56) Thereafter, the parties filed . summary judgment motions. On ·May2, 2016, the Court denied all SU1:Umacyjudgment motions, · with the exception of JHU' s motion for partial summaryjudgment that.JHU' s priority date with respect to the Count is no later than.June :5, 2003, which the Court granted. (See generally D.I. 97, 98) The Court held a bench trial on all remaining ·issues· in. June 2016. (See Transcript, D .I. J 12, 113, 114 ("Tr.")) The parties later briefing (D.I. 108, 110, 1l5,118). and proposed findings . . of fact (D.I.109, 111, 116,.117). - . . . . . . . . Pursuant to FederaLRule .of Civil Procedure52(a), and after having considered the·entire · · . . . . . record in this case and the applicable law, the Court concludes that: (1) JHU has failedto prove that it is entitled to priority ofinvention, and (2).JHU has fa~led to prove that 454's '240' ' application is invalid. The Court's findings of fact and conclusions of law are set forth in detail below. 2 All pending claims of the '690 application (claims 1, 2, 5-17) and the. '240 application (claims 52-66) correspond to the single Count in the Interference. 2 FINDINGS OF FACT . . . This section contains. the,:Court's fmdings .of fact for issues raised qy the parties during trial ··Certain findings of factare also .provided in .connection with .ihe, Court'-s conclusions· of 1aw. A. L ·Patent Applications :at ]ssue ·.Plamtiff.JHU' s '690 application, entitled "Method and Compositions for Detection and Enumeration of Genetic V aiiations," was filed on January 29, .2009.. · (D .I. 22-1 ·Document .1) · The named inventors are Devin Dressman, Hai Yan, Kenneth W. K.iniler, arid Bert . Vogelstein. (Id.) 2. Defendant454's ''240 application, entitled "Bead Emulsion Nucleic Acid·· Amplification," was filed-onFebruary23, 2011. (ATX 1001)3 The named inventors are Gary · Sarkis, Jan Berka,.JOhn Leamon, Kenton Lohman, ·Maithreyan Srinivasan, Yi-Ju Chen, Vinod Makhijatii, Jonathan R9thberg, Steve Lefkowitz, and Michael Weiner. (Id.). The '240 . . application issued as U.S. Patent No. 8,748,102 ("'102 patenf') on.June_ 10,2014 .. (DTX 12)4 3. The '240 application is.a continuation ofU~-s. Patent Application No. 11/982,095, filed on October31, 2007(ATX1005), which is a continuation of U.S. Patent Application No. . . 10/767,899 ('"899 application"), filed on January 28, 2004(ATX1007). The '899 application -claims a number of provisional applications, including u~s. Provisional Patent 3 The Court admitted exhibits from the Interference into evidence (ATX) .. (See D.I. 21-32) The _parties submitted a list correlating exhibit ni.unbers to document descriptions in their joint proposed pretrial order. (DJ. 102 Ex. 8Al) 4 Exhibits submitted by Plaintiff (PTX) and Defendant (DTX) that are cited herein were admitted into evidence. 3 . . Application No. 60/476,592, filed June 6,2003 ('"592 provisional" or."'592 application") (ATX · 1013), and U.S.; Provisional Patent ApplicationNo .. 60/465;071, filed April 23., 2003 ("'071. (-. ·provisional" or"'071.applicatiori") (ATX 1015). The contents of the '071 and ·'592 provisional applications are incorporated by 4. reference in 1heir .entirefy into the "'240 :application. C240 application at 1 :3-6) The '240 application also facorporates by reference co-pendingU.S. -Patent 5. Application No.10/767,779 ('"779.application~'), which' U.S ..PatentNo. 7,323~305 ('"305 patent") on January 29, 2008. (ATX 1001; DTX 12; DTX 13) The '305 patent contains the entire disdosure ofthe ·'592 provisional. (Id.) · .'Procedural History in ;the Patent Office B. · The Board initially accorded JHU a priority date of Ju:~y-S, 2003 and 454 a priority · 6. date ·of June 6, 2003 (the filing date ofthe '592 provisional), making JHU thejunior party and 454 the senior party in the Interference. (See ATX 236 at :3: 16-26) .In the Interference, JHU filed a motion attacking 454's claim to 7. the '592 . provisional's filing date. (D.I. 23-4 at 7.:6-13) The Board denied JHU's motion, ruling that)HU had failed ·to show that the '5 92 provisional did not diselose a reduction to practice within the · scope of the Count. (Id.) 8. In the Interference,.454 filed amotionto obtain the benefit of the '071 application. (ATX 236 at 8:11-16) The Board denied 454.'s motiOn, finding that 454 had failed . . . . . . to establish adequate written description support in the '071 application. (Id. at 1?:24-26) . . . . .Specifically, the Board found that "454 ha[ d] not established (e.g., by citing to data or expert . . . . . testimony)" that use oftestriction enzymes recited in the '071 application "would generate two or 4. , . ·more molecules ofa specific DNA tr8:gin.ent, or more specifically, that an· ordinary artisan would understand thatto bethe case;":as required to.practice step (a) ofthe Count. (Id. at 14:13"'"18) 9. · During .the priori~y ·stage, JHU submitted evidence of priority from theJ anuary to May 2003 time frame; (ATX 3 70 at 5 n.1) .However, the Board found thatthe "evidence cited . . by JHU does not sufficiently.establish thatJHU .conceived ofthe subjectmatter of:Count 1 in the Januaryto May2003 time frame:". (Id.) Specifica1ly,the Board found that the evidence ''[did] not adequately show .JHU ccmceived of elements (a) and (b)" of the Count during that time frame. (Id.) The Board also found that "JHU offer[ed] insufficient non-inventor evidence (testimony or otherwise) to corroborate conception by the JHU inventors at that time." .(Id.) As . . .a result, the Board accorded JHU .a June 5., 2003 conception date. (Id. at 7: 16"'.22) The Board found that JHU reduced the-invention to practice one month 1atet, ·on July 5," 2003. (Id.) 10. With respect to 454, the Board concluded that a preponderance of the evidence showed.and corroborated the fact that the 454 inventors conceived of all elements of the Count before June 2003. (ATX 370 at 17:1-23:9) In particular, the Board found the combination of page 16 of the notebook ofDr. Jan Berka (one of the 454 inventors) (ATX 1094), evidence. regarding experiments conducted between August and December 2002, evidence of experiments conducted in January and February 2003, a "Best Practices" document from February 2003 (ATX .1102; ATX 1103), and an invention disclosure form (ATX 1106,at 107-12), taken as a whole,. established that the 454 inventors .conceived the subject matter of the-Count before JHU's earliest. accorded date; (ATX 370 at 21 :7-23 :2) · 11. ·_The Board held that "as long as the invenfots conceived ofperforming.[step] (a),· i.e., generating .a plurality of molecules ofa particular fragment .of DNA, along with. [steps] (b) 5 through (d), it [did] not matter whether the inventors failed 'to appreciate the vallle of step (a)' ·beyond its use to generate .a control used in the method [as alleged .by JHU] ." (ATX 3 70 at 18:20-19:3) The Board further held that the "evidence establishe[ d] that the inventors appreciated that [step] (a) took place, regardless ofits 'value' in relation to benefits of the protocol in amplifying genomic template DNA." (Id. at 19:3-5) 12. The Board awarded priority of invention to 454, finding that 454 conceived of the invention of the Count before JHU' s conception and was also first to reduce the invention to practice. (ATX 370 at 23:8-9) C. The Court's -Claim Constructions 5 13. The Court construed the term "generating a plurality of molecules of a fragment of deoxyribonucleic acid" in step (a) to mean "generating two or more of the same DNA fragment, not merely generating a plurality of DNA fragments overall." (D.I. 57 at 2) The Court's construction of step (a) does not require any particular method of generating_two or more copies of the same DNA fragment (Levy Tr. at 367:9-19) 6 However, the fragment formed in step (a) is a sequence that is ultimately amplified in the emulsion and sequenced. (Id. at 505:8-17) 14. • The Court determined that the following term required no further construction: 5 In this Opinion, the Court applies the broadest reasonable interpretation of the various terms of representative claim 1 ·of the· '690 application in light of the "originating disclosure." See ULF Bamberg v. Dalvey, 815 F.3d 793, 796 (Fed. Cir. 2016). The Court previously determined that the "originating disclosure" in this case is JHU' s '690 application and its prosecution history, which includes-references to some of 454's patent applications. (See D.l. 56 at 8) 6 The Court refers to testimony given at trial by a particular witness in the following format: "([Witness Last Name] Tr. at [page:line(s)])." 6 "delivering the plurality of molecules of the fragment of deoxyribonucleic acid into _aqueous m.icroreactors in a water-in-oil-emulsion such that a plurality ofaqueous microreactors comprise a single molecule ,of the fragment .of deo~yribonucleic acid, .a single bead capable ofhybridiiing - to the f!agment .of deoxyribonucleic acid, .and regents necessary to ,perform· deoxyribonudeic ·acid amplification." (D.I . .57 at2) There is·no significant distinction between delivering to a microreactor double-stranded DNA s~parately from a bead versus delivering single-stranded DNA pre-hybridized to a. bead. (Levy Tr. .at 405:8-24, 470:14-19; see also Tyagi Tr. at ' 116:13-15} . . 15. The Court construed the terms "deoxyribonucleic acid" and "DNA"to mean "a nudeic acid molecule comprising deoxyribonucleotides." (D.L 57 at 2) · Single~stranded DNA is -a nucleic .acid molecule comprising deoxyribonucleotides. (Levy Tr. at 404:3-9) 16. The 'Court construed the term "a single bead capable of hybridizing to the . ' . . fragment"to mean "a single bead capable of binding to the fragment of deoxyribonucleicacid~" (D.I. 57 at2) The Court's construction of"a single bead capable of hybridizing to the fragment" only requires that a bead be capable of hybridizing to the DNA fragment. (LevyTr. at 390:16-391:14) A bead that has bound to the fragment is capable of binding, and.a bead that has not bound to a fragment may also be capable of binding. (Id.) . 17. ·.The Count may be satisfied-.even ~f the fragment from step (a) is pre-hybridizecl to a bead before deliveryto a.microreactor. (Id. at505:18-506:1) The sequence of the fragment formed in step (a) remains intact regardless ofwhetherit is hybridized to the bead. (Id. at . . ' . . 391:24-393:14) Moreover, when the fragment of DNA hybridizes to the bead, it does not create a new molecule. (Id.) . The DNA fragment and the beadretain their individual identity, even ·_ .7 while hybridiz~.q. (Id. .at 393: 10-14, 459:19-460:4) 18. Hybridization of the DNA fragment :to the bead is through hydrogen bonding, which 'is a non-covalent, electrostatic interaction. . (Id. :at 392 :16-393 :9) Hydrogen bonding can be thought oflike a sock ~ticking to a sweater when it is pulled out of a dryer. (Id.) Justbecause a·sockis.stuckto.a.sweater·does not mean.anew entity.has formed. (Id.) Similarly,just.because a DNA fragment-has hybridized to a bead .does not mean a ·new molecu1e has formed. (Id.) . D. 19. . .Person of Ordinary Skill1n the Art One of ordinary skill in the art at the-pertinent time would have had around four years of research.experience, .a Master's degree or Ph.D. in molecular-biology or other relate~ fields such as .genetics or biochemistry, .and would have been familiar with polymerase chain . reaction ("PCR"). (Tyagi Tr. at 45:15-46:7; Lery Tr. at 353:7-354:4) A person of ordinary skill . '• . ·in the art would have been aware of emulsion PCR as a general concept. (LeryTr.. at 353:25354:4.(statingthat attime ofinvention "there were.a.couple ofve·ry high profile papers·that · describe[d] using.emulsions to either do PCR, in the case of the Hollinger paper, or to use emulsions to.encapsulate other things"); ATX 1099; ATX 1097 af4) . ·K. 20. JHU'sWitnesses JHU calledjust_a"single witness to testify live at trial. Dr. Sanjay Tyagi obtained a B.S. from the University ofRajasthan in India, two M.S. degrees in biology from the Jawaharlal . Nehru University in NewDelhi, and a Ph.D. from the-University of Maryland. (Tyagi Tr. at . 42:16-21; PTX 001) 21. 'Since 1987, Dr.. Tyagi has worked at the Public Health Associate Institute, which . currently is a part of Rutgers University, initially as Associate Professor and currently as a full · 8 Professor.. (Id. at 42:24-43:2) Dr. T:yagi has 29 years of postdoctoral experience in the field of · nucleic aCids, molecu1ar:biology, ·and. cell .biology research. (Id. at43 :5-44:7) 22. .JHU .called ·several .other witnesses to Teading. some. or all of the dec1aration(s) these witnesses submitted as part ofthe Interference proceedings. (See Tr. at 164.:21-175:3; ATX 2024, 2034, 2051, 2079, 2080) 23. JHU presented testimony from six witnesses by declaration: .a. Dr. BertVogelsteinjs a professor of oncology at Johns Hopkins University School ofMedicine, Baltimore, ·MD, and a Howard Hughes Medical Institute investigator. (ATX 2034·if 2) Dr. Vogelstein supervised the work of Dr. Devin Dressman as a post-doctoral fellow in his and Dr.Kenneth Kiniler's laboratory beginning·at the end.of.January2003.and continuing beyond: July 5, 2003. (ld.'if 3) b. Dr. Devin Dressman currentlyworks in research and development for Life Technologies· Corporation in Beverly, MA. (ATX2051 'if2) He is in the "fon Torrent" division at.Life Technologies, which deals with a _sequencing system that .utilizes bead emulsion amplification.for sequencing sample preparation. (Id.) Dr. Dressman worked as a post-doctoral. fellow inthe·laboratory ofDrs. Kenneth Kinzlerand Bert Vogelsteinat _Johns.Hopkins University beginriing at the end of January 2003 and continuing beyond JulyS, 2003. (Id.· if 3) c. · Dr. Hai Yan worked with Dr. Dressman between January2003 and.Juiys, 2003 on.bead emulsion amplification projects. (Id.) d. Di. Kenneth Kinzler. ran a laboratory with Dr. Vogelstein at Johns Hopkins University, as discussed above. (ATX 2034 if 3} e. Ms. Leslie Meszler held the position of Core Manager at the CeJl Imaging 9 Core at:J ohns Hopkins University in 2003 ~ (ATX 2029 if 1) She was responsible for managing thed~y-to-day operations ofthe-Cellimaging-Core during2003. (Id.) One ofherresponsibilities included tracking scientists·' use -of a f1owcytometer. -(Id. ifif 3-:5) She testified ~y declaration) that the flow.cytometer was used byDr. Dressman during the week of April 20, 2003. (Id. ·ir 6) f. -Mr. Jason· Briody is an Assoeiate at Jones Dykstra & Associates, a - specialized services .company that _provides .computer forensics, electronic data discovery, litigation support;-training, and ·computer security services for commercial and governmental clients. (ATX2081 at 6) ·Mr. Briody is primarilyresponsible_forplanningandtechnically executing electronic discovery projects and performing computer forensic analysis. (Id.) Mr. _Briody analyzed files given to him by Johns Hopkins Universit,y for authentication purposes related to this litigation. (Id.: at 4-5) R 454's-Witnesses 24. Dr. Matthew Leyytestified live at trial.· The Court found him to be credible and persuas~ve 25. on everymaterialpoint. Dr. Levy earned a B. S. in-biochemistry and a M. S. in chemistry, both from the - - - · University of California San Diego, ~d a Ph.D. -in molecular -biology from the University of Texas. (DTX.32; Levy Tr. at.347:18-348:7) Dr. Levy is an Associate Professor ofbiochemistry at the Albert Einstein College of Medicine. (DTX 32; Levy Tr. at 348: 10-17) Dr. Levy teaches a variety of classes, including biochemistry, immunology, and chemical biology. (Levy Tr. at . . . . 348:19-21) Amajorfocus of his research is developingnlJ.cleic acid-based therapeutics and diagnostics. (Id. at 349:3-7) He is familiar with genomic DNA isolation, DNA sequencing, and emulsion PCR. (Id. at 349:14-350:1) Dr. Levy has personally conducted emulsion PCR and has 10 experience making.emulsions for.emulsion PCR. (Id. at 349.:18-21) Dr. GaryJ. Sarkis is one ofthe454 inventors. Dr. -Sarkis received a RS. in 26. microbiology .and :an M.S. in biochemistry .and molecular genetics, both from the University-of Pittsburgh. He also received :a Ph.D. in molecular, cellular, and developmentalbiology from the University-ofPittsbu:rgh in 1997. He was a postdoctoral research fellow at Yale Universityfrom 1997 to 2002 and was ~mployed at 454 :as a_research scientist, section leader (sample preparation), training manager, and customer support manager from 2002 to 2006. (Sarkis Tr. at 177:11:..15, 178.:5-179.:4) Hetestified live at trial 27. Janna Lariza Thompson received a B.S. in biology from the University of Vermont and a M.S. in cell artd molecular biology from Central Connecticut State University. She was a research associate from 2001to2006. (Lanza Tr. at 278:3-279:2) She testified live at trial. 28. Dr.Alex de WinterreceivedaB.A. in chemistry from Amherst College and a ' . ' . ' :· ' Ph.D . .in chemistry from.Stanford Univer~ity. He was employed at 454 .as a research scientist from 2001 to.2004. (de Winter Tr. at.322:22-324:9) He testified live at trial. Dr. Jan Berka is one of the-454 inventors. He received a B.S~ and M.S. in .29. . . molecular biology and genetics and a Ph.D. in molecular biology and gen_etics, all from Masaryk .University, Brno, Czech Republic. He was a postdoctoral research fellow at the Barnett Institute · . . . . . . . . . . at Northeastern University in Boston from ~1992 to 1996. ·Dr. Berka was employed at 454 as a senior scientist and director from 2000-2006 .. (ATX 1113 ·irir 2-3) Dr. Berka testified by · declaration. 30. Dr. Maithreyan Srinivasan is one of the 454 inventors. Dr. Srinivasan has a Ph.D. 11' . . in biochemistry: He was .employed at 454 as a prqjectJeader in the Protein :Sciences group from . 2000~2007. 31. (ATX 1116 if~ 2-3} Hetestified by dedaration. ·Mr. Keith :McDade .received .a B.S. in molecular .biology froni the University of Connecticut and a M.S. in computer science from the University of New Haven. He was· employed at 454 ·~s a.research associate from 2000 to 2006. (ATX 1124'~~2-3) Hetestified b,y .declaration. 32.. Dr. John.Leamonis one ofthe 454 inventors. Dr. Leamon received aB.A. in zoology and a Ph~D~ in physiology from the University of Connecticut. He was a postdoctoral· research fellow at the Yale School of Medicine from 1999~2001. ·Dr. Leamon was employed .at · 454 in various positions ranging froni"research scientist to group leader from 2001~2007. (ATX 1114 -~~ 2-3) He testified by .declaration. 33. Dr. Louis ~Ferland received a B.S. in biochemistry from Universite Laval and a . Ph.D. in experimental medicine from McGill University. I-Je was a postdoctoral fellow · .·at the Salk Institute, RegUlatory Biology Department from 1986-1989;. and at the Institut Pasteur, . . . Departement de Genetique Moleculaire du Developpement from 1989.:1991. He was employed at 454 in ·various positions ranging from technical :writer to manager of documentation from 2001~2012. · .34. (ATX 1118 if~ 2-3) He testified by declaration. Mr. William Altman received a B.S. in biology from Guilford College. He was employed at454 from 2001·102013, holding several positions ranging from research assistantt~ senior customer support speCialist. (ATX G. · 35. 1120·~~ 2-3) He testified by declaration. 454's Conception ofthe Invention by December 2002 Dr. Sarkis testified at trial, and previously submitted a deClaration to the Board; 12 indicating th~the recalls discussing the idea for .a method for .analyzing nucleic acid sequences using bead PCR emulsion .amplification With Dr. Berka - which :is ·.reflected in Dr. Berka' s .laboratorynotebookdated June 7.,2002. (Sarkis Tr. .at 180:8-183:5:; ATX 1115if16; ATX .1094 at 16; ATX 1113·'if 18) 36. ·On June 7., 2002, Dr. Berka recorded in his lab notebook the notes ofa conversation with Dr. Sarkis .about the idea ofPCR in water droplets in oil (water-in-oil emulsion) as individual microreactors that would contain a si!1=gle effective copy of a sequence of DNA, a capture.bead, and enoll;gh PCRreaction solution to produce amplified amounts of individual DNA fragments for massively parallel sequencing. (Sarkis Tr~ at 180:8:..183:5; ATX 1115 'if 17.; ATX 1094at16; ATX 1113 'if .18) .. 37.. That same page·ofDr'. Berka's notebook (page 16) also . contains a drawing that depicts the concept ofaPCR reaction occurring from a single head.and a single starting DNA fragment withinthe individualmicroreactors. ·(Sarkis Tr. at 182:7-23; ATX 1115'ifif 17-20; ATX · 1094 .at 16; ATX 11 ll·if 19). The drawing also shows the single-stranded DNA fragment . . . . . . ' attached to a primer on the capture bead· and the resulting double-stranded DNA that would be · present after the amplification process begun. (ATX 1094at16) To the right ofthe-drawing of the microreactor, Dr. Berka noted that "isolated bead bound primer extension" would occur · "inside ofthe 'ind1vidual bead reactors." (Sarkis Tr. at.184:4-13; ATX1115'if 19; ATX 1094 at 16; ATX 1113.if 20) 3 8. Dr. ·Maithreyan Srinivasan testified by declaration that he recalled discussing the idea for a method of analyzing nucleic .acid sequences using bead emulsion amplification with Dr. Berka. (Srinivasan Tr. .at 267:25-268:13; ATX 1116 ·irir 9-15) This idea, also recorded on 13 page 16 of Dr. Berka's notebook (A.TX 1094), was contemporaneously witnessed and signed by Dr. Srinivasan on the _same day it was written downby Dr. Berka: June·7, 2002. (Id.) 3 9. Dr.· Leamon testified ·bydeclarafion about experiments he performed in August 2002 to improve the stabilization of emulsions for bead emulsion PCR. Although Dr. Leamon was able to successfully form the emulsions, after a number ofcyeles the .emulsions were· breaking down or·"crashing:" (Leamon Tr. at 307:2~308:24; ATX 1096at]10-12, 123.; ATX 1114 irir 21-24) On December Tl, ~.2002, Dr. "Leamon proposed the use ofrestriction endonuclease 40. enzymes," known as "4-cutters;":instead of DNaseI, to digest DNA fragments. (Leamon Tr. at 308:25~309:1_7;ATX 1097at1;ATX1l14·ifif25-26) The '592 provisional specifically discloses . . using Sau31, Mspl, and Taql, which are4-cutters. (Lery Tr. at 368:24-369:10; ATX 1013 at 11:24-26) · Dr. Leamon recognized using 4-cutters would make DNA template with at least two .. . . copies ofa fragment of DNA. (ATX J 114 if 25) Dr. Leamon ~uggested using as many 4~cutters as prudent, perhaps 4 or 5, ligating adaptors onto .each pool, and then hybridizing the DNA template onto the beads. (Id.) . 41. On December 19,2002, Drs. Leamon, Sarkis, and Berka attended alectureat Yale University given by.Jennifer Ong. (Sarkis Tr. at 186:18-187:22; ATX 1105 at 93; Leamon Tr. at . . 309:18-25; ATX.1095 af69; ATX 1114"iif 27; Berka Tr. at 303:.1-4; ATX.1113 if 23) Dr.·Ong is one of the co-authors of a well-known journal article entitled, "Directed evolution of polymerase · . . . . . . . . . - - . function by compartmentalized seif-replication;" attributed to Ghadessy-et al., and published in the journal of Proc~edings of the National Academy ofSciences ("PNAS") on April 10, 2001 • ("Ghadessy"). (ATX 1099; Sarkis Tr. at 187:23-188:13; DTX.33) Also on December 19, 2002, 14 ·Dr. -8arkis sequenced test fragments that 454 used as controls and obtained good sequencing. results. (Sarkis 192:19-193:2; ATX 94). -42. Drs ..Sarkis, Leamon, and Betka referenced ihe seminar givell'.at Yale by Dr. Ong in their notebooks. (Sarkis Tr.·at 186:18-187:22; ATX 1105 at 93:; Berka Tr. at 303:1-4; ATX 1095 at 69; ATX l113.if23; Leamon Tr. at309:18-310:8;ATX ll14ifif 27-28) Dr. Leamon :ineluded an excerpt of the Ghadessy ·paper in his lab notebook and noted his beliefthat the emulsion information discussed by Dr. Ong could be used for emu1sion PCR using .a sepharose bead. (Sarkis Tr. at .188:3-l90:l; Leamon Tr..at309:18-310:8;.ATX 1097 at 2-3; ATX 1114. 'ifif 27-28) Dr. Leamon also noted·that·"Andrew Griffith's group has used emulsion based.bead . PCR ·bead capture for translation studies which suggests that the beads maintain their discrete· micelle identity~" an.d included an excerpt from an article b.y Armin Sepp, D'an Tawfik, and. ·Andrew Griffiths ("Sepp"), regarding formation of emulsions, on the next page ofhis notebook ! onDecember19, 2002. (ATX1097.at3-4) The Sepp article is entitled "Microbead display by in vitro compartmentalization s_election for binding using flow cytometry," and was published in Federation ofEuropean Biochemical Societies Letters ("FEBS") in November 2002. (ATX .noo) 43. On December 20, 2002, the day after attending the lecture .at Yale and reviewing . . . ·the Ghadessy and Sepp references, Dr. Leamon performed an experiment and· successfully prepar.ed beads in a water-in-oil.emulsion. (Sarkis Tr. at 190:2-16; ATX 1097 at5-6; Leamon Tr. at 310:9-il; ATX 1114'if 29; Levy Tr. at446:24-447:17) Dr. Leamon included pictri.res l.n his notebook that demonstrate that some of the micelles were suitably-sized for the sepharose beads. (ATX 1114.if 29) . 15 -44. Also .on December 20, 2002, againthe day after attendingthelecture at Yale, Dr. Sarkis performed ;an :emulsion PCR experiment with ··PCR-generated test fragments TF l, TF2, TF3., TF4, TF5., TF6,.F6., aridN7, and$uccessfully.amplifiedthe test fragments. (Sarkis Tr. at 193·:22-195:7; ATX 1105 at 95-96; Levy Tr. at446:24~447::17) Dr.·Sarkis's December 20, 2002 experiment evidenced conception ofthe invention of the· Count, as further explained below. 45. Dr. Sarkis testified that the test fragments· used by 454 originally canie from Curagen Corporation, a sister company to 454. (Sarkis Tr. at 193:3-21) The fragments were 1abe1ed by the well number that they .came from on the microtiter plates; and 454 knew the_ sequence of all of these test fragments. (Id~) The sequence of the F6 test fragment is recorded in Dr. Sarkis' s -lab notebook many times and was used for comparison each time the test fragment · was sequenced in a reaction. (Id.). 46. The F6 test fragment was .PCR amplified by Mr. Altman, a 454 research assistant, who declaration about making .theF6 test fragments. (Altman Tr. at 271:25-27~:20; ATX 1120·'ifif 6-1 O; ATX 1108; ATX 1109) As recorded in his ·notebook, Mr. Altman _ .. successfully sequenced the F6test fragments on July 22, 2002. (Id.;ATX 1109at13) 47. · Ms. Lanza (now Ms~ Thompson) testified that she recorded the sequence of the F6 test fragment, whichwas a PCR~generated fragment commonlyused at 454, in her notebook on · January23, 2003. (Lanza.Tr. at 280:1-20; ATX 1132 at 134; ATX 1126·if ll) The sequence of ' ' F6 is identified on page 134 of her notebook, i.e., ATX 1132 at 134 .. 48. Dr. de Wintertestified that all.ofthe test fragments that were used as controls by · the454 scientists were PCR-generated fragments. (de Winter Tr. at 331:16-18) JHU's expert, _Dr. Tyagi, testified that the test fragments, e.g., F6, would satisfy step (a) of the Count if they 16 were generated ~y PCR. (Tyagi Tr. at 90:5~19) 49. · Dr. de Winter testified thafhe prepared adenovirus DNA libraries for use in the emulsion PCR project and testified about the process. or standard operating procedure that be . . .. . used to prepare the libraries. (de Winter Tr. at 324:6-9, 326::2-327:23.,330: 17-331:4; A.TX 1122 .at 51-52, 65., 69, 71~72, 75-76, 78; ATX 1123 'iii! 10-11) The typical process forpreparingthe library would be to fragment the adenovirus, polish the ends of the fragments. with polymerases or dNTPs, ligate adaptors to the ends of the polished fragments, purify the ·ligation products, capture on streptavidin-coated beads, and then elute the single-stranded DNA fragments. (ATX 1123 ·:ir J 0) SO. Dr. de Winter included in his notebook a detailed sample preparation protocol that he used forpreparingadenovirus templatelibraries for emulsion PCR. (de Winter Tr. at 327:1~23, 330:1-11; ATX 1122 at 60) For this particular protocol, he usually used DNase i to ·fragment the adenovirus. (de Winter Tr. at329:2-15,J30:5:.16; ATX 1122 at59) 51. Dr. de Winter testified that there are a number of ways to fragment DNA, · including using restriction enzymes that recognize particular 4...:base sequences. ·(de Winter Tr. at 331 :24~332:6) 52. Restriction enz;yme digestion can be used to generate overlapping genomic· sequences for sequencing because they-will eµ.ch cut at a different site. (Tyagi Tr. at 125:15-19; . Levy Tr. at 388:9-25) As a result, if a genome is cut with one enzyme it will generate a particular set of fragments, and if it is then cut with .a different enzyme, it will generate a different set of overlapping frigments. (Tyagi Tr: at 125:20-127:3; Levy Tr. at 388:15-25) 53. Both Dr. de Winter and Dr. Sarkis testified that, depend1ng on the experiment, 17 they would sometimes perform emulsion PCR of the test fragments alone, and other times they would mix test fragments together with adenovirus fragments. (Sarkis Tr. at 195:19-.196:1; d_e Winter Tr. at 332:7-333-:7) 54. Dr. Sarkis testified at trial that the 454 inventors wanted clonal amplification, so they designed their experiments with the goal of having a single effective copy per-bead. (Sarkis Tr. at 182:24-183:21) Clonal amplification means amplifying a single DNA fragment in isolation from all others. (Sarkis Tr. 55. at217:5~23) After December 2002, 454 scientists continued to perform experiments to optimize the conditions and develop a product that could be easily used by their customers. (Sarkis Tr. at 195: 8-18) H. The Poisson Distribution 56. The "Poisson distribution" is a statistical tool that can be used to predict the number of times that a given event occurs within a certain interval or physical space and is well~known and widely accepted. (Levy Tr. at 358:2-15) 454's expert, Dr. Tyagi, testified that the "Poisson distribution is [a] statistical distribution, basically, that scientists use when they want to deliver single cells into a well or single molecules in a reactor and so on." (Tyagi Tr. at 98:17-24) Using the Poisson distribution, it is possible to estimate the number ofbeads in a population that will be attached to more than one DNA fragment, to a single DNA fragment, and to no DNA.fragments. (See Levy Tr. at 358:11-15) · 57. The Poisson distribution has been repeatedly validated through empirical testing and experimental verification. (Id. at 361:4-15; 1Yagi Tr. at 133:9-23) Dr. Tyagi explained: "The Poisson distribution stands on its own. It's always true," and "Poisson statistics are always 18 true, nobody is denying thaC' (l;yagi Tr. at 137::2-3, 138:25-139:1) The Poisson distributionis reliable . evidence· ofwhat :actuaHy transpired during the. experiments performed by 454's scientists. (Lery Tr. at 361:16-22, 498:13-24) The number ofcompartments .present in a 454 emulsion .PCR reaction can be .calculated by :kn_owing the diameter of the compartments and the volume ofthe aqueous solution. (Id. at .365:11-22) Knowing the number of compartments allows·oneto]mow if enough compartments.have been,generatedto encapsulate all of the beads and all oftheDNAfragments in the experimenta1 setup. (Id.) . 58. . The Poisson distribution can be used to predict whetherthe emulsion PCR ·experiments performed b.y 454. contained some population of beads that ·had on~y-a ·single DNA· fragment attached . .(Id. at 358:9-15) .The December 20, 2002 experiment and each subsequent emulsion PCRexperiment discussed hereinafter would he expected to conform· to the Poisson distribution. (Jd; at 359:23-360:11) Therefore, based on 454's experimental setups and the Poisson distribution, one can reliably conclude that those experiments included more 'than one microreactor with a single bead and the same single DNA fragment. (Id. at 365:4-366: 15) . 59. The sequencing data obtained from the 454 experiments further supports the finding that the Poisson distribution applies to those experiments. ( 479:20~480:20, 481:24-482:6, 487 :5-18, 490: 18~24, 496:3-12) The 454 emulsion PCR experiments produced clear sequencing data showing individual sequencing reads were obtained. (Id at 361 :23-362:13, 363:5-:-13; see also ATX 110~ at 11) This means there must have.been apopu1ation o.femulsions in the experimentthat had a single DNAfragment and a single bead. (Levy Tr. at360:18-361:3, 361:23~--362:13,490:25-4.91:10) Drs. Sarkis and de Winter testified that their experiments . . . produced multiple microreactors containing one bead with a single unique copy of the DNA · 19 fragment, because clear sequencing results would not have been obtained if the microreactors. contained multiple different fragments. (Sarkis Tr. :at 240: 10-24.1::24, 248:9~249:1; de Winter Tr. at .332:1~333::7., .345.:13-19) :-.Clear-sequencing tesrilts were foundin the .experiments performed after Dec_ember 20, 2002, induding experiments that mixed test fragments with adenov1rus fragments. (See, e;g.~·:Sarkis Tr. .at 195:19-196:17.,212:12-213:11) 60. If there was more than :a·-single fragment on a bead, then the sequencing data would not be dear--'- it would be mll4dled and would not map to a known -sequence. (Lev,y Tr. at 362:14-20) If one can obtairi sequencing data, that means there was only one sequence onthe . bead. (Tyagi Tr. at 546:6-8) Sequencing data also would not be produced if there were more than one bead per compartment. (Levy Tr. at490:25-491:10) I. -454 ·Experiments :W.ere ·Reductions to '.Practice 61. · 454 scientists performed a number of experiments betweefl: January and March of 2003; each of which qualifies as a reduction to practice, as explained.below. 62. OnJanuary 14-15, 2003~ Dr. Sarkis ·setup emulsion PCR reactions using a combination ofadenovirus DNA.and test fragments (TF4, TF6, TF7, and F6)·with an input of 600,000 beads and 1.2 million molecules of DNA (two copies per bead) and successfully obtained sequencing-results-for the F6 test fragment from emulsio:ri-PCR-generated beads. (Sarkis Tr. at 199:8~2Q0:22; ATX 1105at112-13) Using this setup, based on a Poisson distribution, at leasttwo niicroreactors would have contained a bead and a copy of t~e same F6 fragment. (Sarkis Tr. at 200: 15-22; see also Levy Tr. at 359:23.J60: 11} . - 63. '. . . . . _On January 22-23, 2003, Dr. Sarkis set up multiple emulsion amplification experiments on test fragments with a goal of creating microreactors containing one copy of DNA 20 fragment per head. (See "Sarkis Tr. at202:l5-203 :13.; ATX 1105 at 118) After January 23, 2003, his team generally used two. copies per bead as input. {Sarkis Tr. at 204:7-.10) 0ne of the 0 J anua:cy 23 .experiments· 'Showed ·successful amplification us~g-:emu1sion PCR on a mixture of severaldifferent test fragments (TFl, TF2, TF3, tF4, TFS., TF6, TF7 and N7). (Id. at · 203.:14-204:18; ATX 1105at119) At least two microreactors in this experiment would have· :contained a single bead and a single copy oftest fragment. ·(ATX 1105 at 119.; see also Levy Tr. at359:23-J60:11) Dr. Sarkis testified that, with respect to this experiment, it was likely that.·"not only two, but thousands" of the microreactors would have: contained a single'bead and a single copy of the same DNA fragment, based on the Poisson distribution. (Sarkis Tr. at 204 at 12-18) 64. On January 28-31,2003, Dr. Sarkis set up emulsion PCRreactions using a combination of adenovirus _DNA .and ·p6 test fragments with an input of 0.1, 2, and 10 .copies per · bead and successfully obtained sequencing results for the F6 test fragment from ~mulsion-PCR- ,generated beads. (Sarkis Tr.,at 207:13-20.8:14; ATX 1105at141-42) Using this setup, and based on a Poisson.distribution, at least twomicroreactors would have contained a single.bead . . .and a single copy of the same F6 fragment. (ATX 1l15-if29; see also Levy Tr. at . 359:23-360: 11) These experiments were corroborated by another 454 scientist, Ms. . Lanza~ who referred to this experiment in her notebook on January 30, 2003 and included in .it a copy ofthe. same library distribution map found on page 143 ofDr. Sarkis.'s lab notebook. (Sarkis Tr. at 208:4~209:15; Lanz~Tr. at285:4-286:12; A.TX 1105 at 143; ATX 1125 at 9; ATX 1126if13) 65. Ori. February5, 2003, Dr. Sarkis performed emulsion PCR and obtained positive sequencing results from samples made by mixing 600,000beads and 1,200,000 fragments of adenovirus DNA with linkers (an input ratio of two copies per bead). (Sarkis .Tr. at . 21 · 209:24-210:25; ATX 1106at1-2)_ The sample also contained F6 test fragments as a control. (ATX 1106 at l-2;·LevyTr.~at358:16-359:12) Based.on:a.Poisson.distribution, at1easttwo microreactors would have contained a single bead and a -sir~g1e copy ofthe -sam·e fragment. _(Sarkis Tr. at 210.:19-25) Dr. Levy testified that, the Poisson distribution, about 27% of the microreactors :in this experiment would have had asingle copy of the same DNA fragment attached .to a single·bead. (Levy Tr. at 358:16-359:22; see also Sarkis Tr. at 210:19-25) 66. On February 6, 2003, Dr. Sarkis performed emulsion PCRand obtained positive sequencing results from samples containing adenovirus DNA library mixed with TF6 test . . ·fragments.· (Sarkis Tr. at 211:1-212:11; ATX 1106 at 11-J2) ·Based on a Poisson distribution and the setup of this experiment, .at least two microreactors would have contained a single bead .and a single copy of the same fragment. (Id.·; see also 67. ~evy Tr. at 359:23-360:11) · · On February 7, 2003, Dr. Sarkis performed two.emulsion PCR reactions and obtained positive sequenCing results from samples containing beads mixed with adenovirus DNA library mixed with TF6 test fragments at a ratio of two .copies per bead. (Sarkis Tr. at 212:12-214:8; ATX 1106 at 13-14, 19-20). Based on a Poisson distribution, at least two . . ·microreactors would have contained a single bead and a single copy of the same fragment in these experiments. (Sarkis Tr. at 213.:5-11, 214:3-8; see also Levy Tr. at359:23-360:11). 68. .On February 10, 2003., Dr. Berka recorded in his lab notebook the notes from a meeting called by Kent Lohman, at which_ information on the progress of emulsion PCR was presented. (Berka Tr~ at304:11-17; ATX 1095 at 107; ATX 1113 ·ir 26) Dr. Berka·noted that emulsion PCRyields about 10 million copies per bead and indicated "IT IS A GO!" (ATX 1095 at _107) 22 ' ·69. . At this point, Dr. Sarkis believed he'had optimized conditions for bead emulsion PCR and began preparing a documentrelating to "Best Practices" for "Pol.ymerase Emulsion . ' . :Chain Reaction." (Sarkis Tr. at 214:9-216::13;.ATX 1102;ATX 1103; ATX 1115 ·~~35-38) The purpose of the Best Practices document was to standardize methods at.454 for emu1sion PCR. '(ATX 111-S '~ 35) Dr.. Sarkis prepared the first draft w:lth Dr. Ferland, who edited it on _February 12-13, 2003.. (Id.:; Ferland Tr; at 268: 14-271:22; ATX 1118 ·ifif 2-5, 11~15; ATX 1102; ATX 1103) The -"Summary" section of the Best Practices document describes, inter alia, the 70. .six main steps_ofbead.emulsion PCR: (l)template quality control, (2) PCR solution preparation, (3) binding of the item fragments to the DNA capture beads, (4) emulsion preparation, (5) amplification, and (6).recovery of the DNA template carrying beads from the emulsion. · . (Sarkis Tr. at 216:14~217:23; ATX 1102; ATX 1103) The summary further notes that "the . . . . . . . ·emulsion format ensures the physical separation of the beads into 100 to 200 µm 'microreactors' -within this single tube, tlms allowing for clonal amplification of the template fragments." (ATX · 1102 at 2) The "Purpose" section ofthe Best Practices document repeats the concept of 71. clonal amplification wherein ''[ sJingle copies of the template species are hybridized to DNA capture beads, resuspended into complete PCR Amplification solution, and emulsified into microreactors (100 to·200 µmin.diameter), after which PCRamplification ·generates 107--fold amplification of the inttialtemplate spedes." (Sarkis Tr. at217:24_;219:8; ATX 1102; ATX. 1103) 72~ · On February 19, 2003, Dt. Sarkis performed two emulsion PCR experiments, and 23 obtained positive sequencing results from samples containing-beads mixed with adenovinis DNA library fragments :and PCR-:generated test fragments TF6 :and -F6 at .an :input ratio· of one copy per bead. (Sarkis Tr..at219:9-220:10, 220:21-221:14; ATXJ 106 at79, 84-'85) ·Based onaPoisson distribution, at Ieast two 1riicroreactors would have contained a single :bead and a single copy of the same fragment during these experiments. (Sarkis Tr. at219:24-220:4, 221:10-14; see.also Levy Tr. at 359:23-360:11) 73. On February 27, 2003, Dr. Sarkis inserted into .his notebook an invention diselosure document entitled "Clonal DNA amplification and immobilization on .emulsified niicroparticles." (Sarkis Tr. at 222:24-225:3; ATX 1106 at 107-12) The invention diselosure identifi~s Dr. Berka's notf'.book (ATX 16) as the conception of the invention on June 7, · 2002, and references reductions to practice based on, inter alia, documents dated December 19:--20, 2002(ATX1097 at 2-6); December31,2002 (ATX 7-9); January14, 2003 (ATX 1105 at.112); February-4-8, 2003 (ATX1105 at 147, 151-52; ATX 1106at1-45); and the· · February 12, 2003 Best Practices document (ATX 1102). The "Prior Art" section of the invention disclosure also identifies the Ghades~y and Sepp papers. (ATX 111) 74. On March 7, 2003, Dr. Sarkis obtained successful results from emulsion PCR · conducted on samples of mixed test fragments and single test fragments. (Sarkis .. 221: 15-222:23.; ATX 1106 at 148-50) These experiments were set up at varying input ratios of 0.1, 0.5; 1, and 2 copies per bead. (ATX 1106 at 148) Based on a Poisson distribution, at least two microreactors would have contained a single bead and a single copy of the same fragment. (Sarkis Tr. at 222:16.,.23; see alsoLevy Tr. at 359:23-360:11) 75. On March 28, 2003, Dr. de Winter performed emulsion PCR and obtained 24 positive sequencing results from samples containing _only a m1xture oftest fragments TF3, TF4, - TF5, and TF7 .at input ratios of O.Ql., 0.1, and 1 copies per bead. (Sarkis Tr. at 225;4-227:6; de Winter Tr. .at .3.34:16-19, 335:2-337:13:; ATX 1122 a~ 84~85) Based on .a Poisson distribution, at least two rilicroreactors would have contained .a single bead and a single copy of the same fntgment. (Sarkis Tr. at226:25-227:6; de Winter Tr. .at337:5-13-; see also Levy Tr. at .359:23-360:11) to -Practice .J. ;Corroboration ofthe Above-Described Reductions 76. Drs. Sarkis, Leamon, Berka and the othet'inventors worked collaboratively with each other and With other454 emplqyees ~on includir~g Ms. Lanza, Dr. de Winter, and Mr. McDade bead emulsionPCR. (Sarkis 179:5-9, 180:20-181:17, 185.:6-186:8, 205:14~206:9, · : 208:4-209:15, 225:4-226:4; Lanza Tr. at278:25-279:6, 280:21-288.:9; see also ATX 1117, 1118, 1119, 1120, 1121, 1123, 1124, 1126) · 77. Mr. McDade testified via declaration that he recalled discussing with Drs. Berka, Sarkis, and Leamon their idea Jor a method for analyzing nucleic acid sequences using bead emuision PCR. (McDade 264:14-267:2; ATX 1124·'ifif 10-13) Mr. McDade corroborated Dr. Sarkis's testimony, as well as the testimony ofDrs. Berka and Leamon(via declaration), regarding the discussion recorded on page 16 of Dr. Berka's notebook (ATX 1094) onJune 7, 2002. (ATX 1124 ifif 10-13) 78. Mr. McDade also corroborated experiments conducted by Dr. Leamon from August to December 2002, and specifically recalled that Dr. Sarkis performed a bead emulsion PCR experiment that resulted in template amplification and sequenceable product just before . Christmas in December 2002. (McDade Tr. at267:3;..267:18; ATX 1124.'ifif 14-15) Mr. McDade . · 25 a1so corroborated experiments conducted by Dr. Sarkis from January to February 2003 and recalled Dr. Sarkis creating the Best Practices document for bead .emulsion _PCR around .that time. (McDade Tr. at267:19"'."24; ATXJ 124ifif16.:17) 79.. Dr. Berka testified by dedaration that in the 2002 to 2003 periodjt was ·coinmon practice in the ,group tO conduct fonnal and informal meetings to discuss various aspects of the projects that were ongoing; it was his practice to keep brief notes of the topics discussed at the meetings. (Berka Tr. at 302:16-20; ATX 1113··if22) Dr. Berka testified about severa1 meetings that occurred in January and February 2003 relating to the emulsion PCR project and identified the peop1e in the _group from ·various sections who would have attended such meetings. (Berka Tr. at303:5"".306:16; ATX 1113'ifif24-30; ATX 1095 at 85, 90, 107~08) 80. Dr. Sarkis testified thafbefore the 454 inventors came up with the idea for· emulsionPCR, 454 was working on an alternative approach called PT-PCR ("pico-titer poiymerase chain reaction"), in which m_icroreactors were formed inside tiny wells on glass pico-titer plates rather than using water-in-oil emulsions. (Sarkis Tr. at 185:6-186:2; Lanza Tr. at 280:21-281: 11) Ms. Lanza was working on PT-PCR amplification, and - because the two technologies had a lot of commonalities - she and Dr. Sarkis often collaborated. (Sarkis Tr. at 185:6-186:2) As Dr. Sarkis explained, he and Ms. Lanza were "sort ofracing to find the·right solution." (Sarkis Tr. atJ86:1-2) 81. .Ms. Lanza testified at trial regarding the almost daily collaboration she had with Dr.· Sarkis and others about the emulsion PCR project in the J an11ary to February 2003 period. . · (Lanza Tr~ at 280:21-288:9) Because they were comparing results; Ms. Lanza was aware of Dr. Sarkis' s experiments, and in particular that he was performing emulsion PCR using adenovirus 26 libranes and F6 test fragments using low input ratios. (Id. af288: 10-289:1, 282: 17-283:1) 82. ·Ms. Lanza testified thcit, ~y February 2003, everyone at 454 was.generally .aware -of the _success of emulsion PCR.. (Id. at 288:10~289: 16) Ms. Lanza noted that an .experiment she . was conducting on february 14, 2003 might have been her final PT:..PCR run. (Id.:; ATX 1125 at 37) Ten .days 1ater, on February 24, 2003, Ms. Lanza included a summary statement in .her ·notebook indicating that -'~given [the] recent. success of EPCR and PTPCR' s contamination · issues,. all efforts Will be on EPCR from now on." (ATX 1125 at 70) 83. Ms. Lanza 'began conducting .experiments in March 2003 using the recipes for the emulsion oil and PCR mix from the EPCR Best Practices document prepared by Dr. Sarkis, which she pasted into her notebook. (Lanza Tr. at 289:17-290:15; ATX 1125 at 108-13, 116-17) 84. . Dr. de Winter also ran emulsion PCR experiments using the procedure provided in the Best Practices document. (de Winter Tr. at 333:8-334:3; ATX 1122 at 74; ATX 1.103) On February 18, 2003, Dr. de Winter inserted a graph in his notebook showing the ainount of adenovirus sequenced with various samples, comparing Dr. de Winter;s conditions, Dr. Sarkis's conditions, and Karrie Tartaro's (another 454 emplqyee) conditions. (ATX 1122 at 74) The graph on page 74 of Dr. de Winter's notebook (ATX 1122) is the same a~ the graph on page 64 of.Dr. Sarkis's notebook (ATX 1106) because Dr. Sarkis and Dr. de Winter were sharing. -information. (Id.) 85. The testimony of Dr. Ferland (ATXl 118), who worked on the Best Practices document; corroborates the testimony of Dr. Sarkisthat he had conceived and reduced the ·fuvention to practice prior to February 12, 2003. ·. (ATX 1102; ATX 1103) ·86. The Invention Disclosure document and the documents referenced therein 27 ·corroborate a complete and clear conception andreductionto practice of the invention of the Count. ,(ATX 1097 at.2-6; ATX 1102;.-A.TX 1103.; ATX 1105 .at 112, 118-34, 147, 151-52; ATX - 1106 at1-45; ATX 1131) 87. 454 demonstrated diligence .at least from January 15, 2003 through 454's construcfive reduction to practice ori June 6, .2003. (ATX 252, 1 113, 1114, 1115, 1116., 111 7, 1118, 1-119, 1120, 1121, 1123., 1126) ·K. The '.592 Application Describes .and Emibles 1he ·Method of the Count 88. The ·'592 provisional .application describes and enables at least a single .embodiment that falls within the scope of the Count. (Levy Tr. at 394:2-10) 89. The '592 provisional discloses the preamble of the Courit. (ATX .1013.at1 :14-19) 1~ 90. The '.592 Provisional Describes .and Enables Step (a) Use of a Type II restriction em;yme will generate two or more. copies of the same fragment when used to digestmtiltiple copies of aDNA target. .. (Tyagi Tr. at 82:14-83:2, 1iS:l1-16; Levy Tr. .at.368:17~369:10) Notably, the '592 provisional states: Suitable methods include ... digestion with-one or more restriction endonucleases (RE) to generate fragments of a desired · range of lengths from an initial population of nucleic acid , molecules~ Preferably, one or more of the restriction enzymes have distinct four-base recognition sequences. Examples of such e~ymes. include, e.g., Sau3Al, MspJ, and Taql. . . . In other embodiments, the restriction enzyme is used with a type IIS .restriction enzyme~ (ATX 1013 at 11 :20-12:2) . · 91. . - . . A person of ordinary skill in the art would know that Type II restriction enzym~s will produce the same set each time they are used to digest DNA. (Levy Tr. at 28 369:23~371 :12;DTX 1at48 ("When a DNA sample is treated with one ofthese fl)pe II] -enzymes, the same set offragments is always produced,.assuming that all oftherecognition sites are cleaved.")) Dr. Tyagitestified that when a Type Il restriction enzyme is used io conduct a · complete .digestion, the same set of fragments will always be obtained. (Tyagi Tr.. at 123 :18-25) 92. When using restriction endonuc1eases for DNA :digestion, standardpractice in the field is to conduct a:complete digestion, so that all ofthe available restriction .endonudease recognition sites have been cleaved. (Levy Tr. at371:17-372:10) Dr. Tyagiagrees the default use for restriction enzymes is to do a complete digestion. ('T.yagi Tr. at ·123.:2-4) Thus, when a reference calls for a restriction .endonuclease digestion, a person of ordinary .skill would have understood that the DNA target should be digested to .completion. (Levy Tr. at 3 72:7-10) 93. The-'592 provisional does not.contain any.statements.that suggest a partial digestion, i.e., a digestion that is not to completion, should be conducted. (Levy Tr. at 372:11-20) Dr. Tyagi agrees that nothing in the454 applications says to do 9nly a partial digestion with a Type II enzyme. (Tyagi Tr. at 125:11-14) 94. Given their common use in the field, a person of ordinary skill in the art would have understood how to use restriction enzymes to digest genomic DNA even without expliCit guidance as to the protocol. (Levy Tr. at 371 :9-16) Moreover, Dr. Tyagi agrees Type II restriction enzymes come with instructions explaining their use. (Tyagi Tr. at 122:6-14) 95. · A person of ordinary skill in the art would not have used the conditions in Example l of the '592 provisional with a restriction enzyme. (Levy Tr. at 383:19-384:11) A person of ordinary skill would have known that restriction enzymes and DN ase I are very different enzymes that require different conditions. (Id.) 29 96. The "'592 provisionalis-Clearthatthe.templatenucleiC acid can be constnicted from anysource.ofnucleic.acid,·inc1udingtissue. ·(LevyTr. at371:3-10;ATX 1013at11:20-21 ("The template nucleic .acid can be .constructed from .~y .source. of nucleic acid, ~.g.; any cell, . . tissue, or· organism .... ")) A person· of ordinary· skill would have known that tissue is composed of more than one cell, which means it contains more than one copy of.a genome. (Levy 373:17-19) Ifrestriction digestion was conducted on DNA isolated from a single diploid cell, .which contains two copies of every gene, then two copies of the same DNA fragment would also ·be generated. (Id. at 381:15-25) 97. The '5.92provisional also explains.that the DNA can.originate from a · . single"".celled organism like a bacteria or virus. (Id. at 373:20-374:19; ATX 1013 at 8:12-13 ("DNA may be derived from any source, including .... bacteria or [a] virus.")) A person of · ordinary skill would have known that it is common laboratory practice to isolate bacterial or viral genomic DNA from a cell culture .. (Levy Tr. at 374:20-377:7; DTX 18) Indeed, commonly available laboratory protocols isolate bacterial DNA ·from a culture of cells. (Id.) Those cultures contain trillions of cells (DTX 18 at2.4.5 (stating 100 mL culture will have 108 to 109 cells/mL)), . . which means there will be trillions of copies of the.genome available for isolation, and subsequent ·restriction digestion. (Levy Tr. at 3? 6: 13-3 77.:2) · 98. The '592 provisional states: "Template libraries can be made by generating a complementary DNA(cDNA) library from RNA, e.g., messenger RNA (mRNA)." (ATX 1013 at 12:3-4) Dr. Tyagit~stifiedthat converting mRNA to cDNAwill result in the formation of multiple copies of the cDNA if multiple copies of the mRNA are present. (Tyagi Tr. at 99. 127:4~ 7) Itiswell-known to those of ordinary skill in the art that most mRNAs in a cell 30 exisfinmultiple copies. (Levy 378:10- 379:5; DTX 11 at 107 ("Thousands of RNA transcripts -can 'be ·made from. the same DNA segment during each .cell .generation:")) Each mRNA-.can :ser\re as -a template for reverse.transcription, which converts RNA into-DNA. (Levy Tr. at 378:17-24, 381:2-4) The result is that multiple copies of an identical DNA m~y-be _generated, even when DNA is isolatedfroma single cell. (Id. -at 378:2-9., 381:2-14) · 100. In .addition, cDNA libraries are typical~y _generated b.y using R T..:PcR. (Id. at 378:17-379:5) The'.592 provis_ional describes the use ofRT-PCR to convert RNAtoDNA. (Id. _at.380:11-23; ATX 1013 .at 64:6-8 ("While DNA is the preferred template, RNA and PNA may ·be converted to DNA by known techniques such as random primed PCR, rever~e transcription, RT-PCR, or a combination ofthese techniques~")) RT-PCR is a process· where reverse transcription is coupled to a PCR amplification step, which will result in the formation of millions of copies of the DNA. (Lery Tr. at 380:24-381 :8) ·Using this process, even if there is only a singlemR1'-fApresent ina cell, millions of copies of DNA would be generated. (Id. at . . . . .381:9-14) Dr. T:yagi testified that use ofRT-PCR will generate multiple copies of a.cDNAfrom a singlemRNA. (Tyagi Tr. at127:8-14) 10 l. PCR or RT-PCR amplification of DNA is a method of fragmenting permitted by step (a) of the Count. (Levy Tr. at .503 :1-21) Indeed, that is the method of fragmenting disclosed .in the '690 application._ (Id.) Moreover, ifthe PCR or RT-PCR amplification products are subsequently fragmented with restriction enzymes; this would generate two or more copies ·of the · same DNA fragment. (Id 465:11-466:1) · 2. . 102. The.'592-Provisional Describes.and Enables Step (b) The '592provisional explains that the capture of beads andDNA in 31 microemulsions will follow the .Poisson distribution, and result in a subset of emulsions with .single beads hybridized to single DNA fragments. (Levy Tr. at 389:25-390:15; ATX 1013 at· .21::16:-19) ·103. The .,592 provisionaLc~ntains an extensive discussion on how:to make and use emulsions. (ATX 1013 at29:28-31:4, 85:25-86:18) · 104. A person of ordinary skill in the art would have understood the '592 _provisiona1 to teach that DNA can be harvested from almost any source, and would not ·be limited to only a single cell. (Levy Tr 372:21-373:16) Dr. 1yagi' s opiriion that the '592 provisionalteaches that only·"a cell" would be-used as a starting source of materia1 for step (a) ofthe CounOgnores that the same quoted sentence from the '592 provisional teaches that a template can be. harveste4 from tissue or organisms. (Tyagi Tr. at 75:_16-76:8;Levy Tr. at 377:14-378:1} .3. · 105. e~g., The '592 Provisional Describes and Enables Steps (c) and (d) · The '592 provisional discloses the. amplification step covered py step (c). (See, A'TX 1013 at30:8-13, 31:6-29) 106. The '592 provisional discloses detecting amplified copies, as described in step. (d). (See, e.g., id. at 32:15-27) L. The ·'071 Application Describes and Enables the Method ofthe· Count 107. The '071 provisional describes and enables an embodiment withinthe scope of the Count. (Levy Tr. at 407:3~10) J 08. . The ·'071 provisional satisfies the preamble of the Count by describing methods of sequencing. (Id. at 407: 11-408:3; ATX 1015 .at 3 8 (pyrophosphate sequencing)) 32 , 1. 109. The '071 Provisional Describes and Enables Step '(a) A person of ordinary skill would have understood that the -, 071 provisional describes and enables step (a) of the Count. (Levy Tr. at 409:5-19) In particular, the '071 provisional states at page 40: "DNA isolation prior to sequencing is performed by any of several commercially available methods. Fragmentation can be performed by one of many methods, including physical and sonic shearing, DNA restriction endonuclease digestion, and nuclease treatment." (ATX 1015 at 40) 110. Dr. Tyagi agreed that the language on page 40 of the '071 provisional could describe step (a). (Tyagi Tr. at 531:3-11) 111. All commercially available kits for DNA isolation at_ the time the '071 provisional was filed were designed to isolate DNA from bulk samples, e.g., tissue samples or cell cultures. (Levy Tr. at 409:20-410:10; Tyagi Tr. at 553:11-21; DTX 16; DTX 20; DTX21) There were no well-known, commercially available kits for the isolation of DNA from a single cell at the time the '071 provisional was filed. (Levy Tr. at 413:25-414:4) Thus, a person of ordinary skill would have understood that the '071 provisional describes preparing DNA samples with multiple copies of a given genome. (Id. at 418: 11-20) 112. The '071 provisional does not disclose any methods or techniques for isolating DNA from a single cell. (Id. at 414:5-8) A person of ordinary skill would not have understood from the '071 provisional that DNA should be isolated from a single cell. (Id. at 414:9-13) Indeed, because it teaches the use of commercially available kits, the '071 provisional suggests the opposite. (Id.) 113. Isolating DNA from a single cell has disadvantages associated with it. (Id. at 33 \_\._ 414:14-415:2} For example, becausethere were then no commerciallyav.ailable kits to do it, one would ·have to design .a protocol to isolate the DNA. (Id.) In :addition, there is on1y one copy of the.gen<?me·available, leaving a verylimitedamount ofmateria1 withwhichto work. (Id.) Ifany · DNA were lost, it co:rild .not-be sequenced. (Id.) ·For these reasons, a person of ordinary skill in the artwould not work with a single cell unless she had to do so. -(Id.) 114. The '071 provisional also de~cribes fragmenting DNA With restriction enzymes. . . (Id. at 415 :3-19; ATX 1015 at 40) The use of restriction enzymes to fragment DNA isolated with commerciall.y available methods results in th~ :generation oftwo or more identical DNA fragments. (Levy Tr; at 415:12-19) 115. A person of ordinary skill in the art would have understood reference to restriction · a reference to Type II restriction enzymes. (Id. at 415:24-416:13) Both Drs. Levy and Tyagi testified that Type II restriction enzymes were the only commercially available restriction enzymes when the '071 provisional was filed. (LeVy Tr. at416:4-13; Iyagi Tr. at · 550 :21-23) Type II restriction enzymes were also the oril.y type of restriction enzymes used in molecular biology laboratory applications. (Levy Tr. at 369: 17-22; DTX 1 at48 ("The . importance of the zype II restriction endonucleases for gene cloning cannot be overstated.")) Dr. •Tyagi testified that a person of ordinary skill would have used Type II restriction enzymes to . . . digest DNA. (Tyagi Tr. at 121.:5-8) 116. The '071 provisional also descri_bes generating template for SNP analysis via PCR. (Levy Tr. at417:12-418:10; ATX 1015 at 76 ("Using primers targeting sequences in close . - proximity to known SNP-containing regions of interest on chromosome 21, we can amplify individual, large fragments and load them onto our capture beads in Eppendorf tribes .... ")) 34 PCR amplificationgenerates millions of copies of.that fragment. .(Levy Tr. at 418:8-10) 2. 117. The '071 Provisional .n~scribes :and Enables 'Step .(b) A person of ordinary skill would have understood that the ·-'071 provisional .describes.and enables step (b) of the Count. (Id. at41R:2J-419:3) For exainple,the '071 provisional states: A second approach to amplifying and capturing.both strands will be to amplify the fragment library offline in a single tube using oil and surfactant-~based .emulsions to encapsulate the capture ·beads, template and PCR reaction·mix. Thisapproachwill maintain the clonalify of the amplification . . . .. The average size of the emulsion. capsules must be optimized to maximize the _·.·number of single beads containing single strands of DNA, that can be incorporated within a single emulsion volume. An adequate volume-tO-bead ratio must be maintained in order to insure a maximum number [of] single bead capsules. (ATX 1015 at 46-47) 118. The paragraph quoted.above (from pages 46to 47 ofthe·'071 provisional) describes emulsion PCR. (Leyy Tr. at 419:24-420:2) It also describes using the fragment library that was generated according to the methods described on page 40, i.e., step (a), as template. (Id. at 420:9-15) It further describes all of the requirements of step (b ), i.e~, using emulsions to capture beads, template, and PCR mix. (Id. at 420:16-20) 119. The '071 provisional explains that there should be a single DNA fragment per emulsion. (Id. at420:21-421:13,422:4-8; ATX-1015 at 46-47 ("This approach will maintain the clonality of amplification . . . . [Emulsions] must be optimized to maximize the number of single beads containing single strands of DNA, that-can be incorporated within a single emulsion volume~")) 35 120. The '071 provisional provides guidance on the parameters that need to be .considered in ·setting up the emulsion PCR. {Levy Tr. .at 421:14-19). In particular, it notes the .average .size ofthe capsules neede~ to make ·sure there isa single bead and a single template per emulsion. (Id. at421:20-422:8; ATX 1015 at46-47) With these disC1osures'in mind, a person of qrdinary skill in the art would ~have ·looked to literature available at the time for .details on how to optimize:the.emu1sion size. (Levy Tr. at422:9-:16) 121. Tawfik and Griffiths published an article in Nature Biotechnology~ a well-known journal to persons of ordinary skill in the art -tha(involves making emulsions forthe encapsulation of biological reactions. (Id. at 422:17-423:19; DTX 23) The publication contains· detailed .guidance on:how to form the emulsion compartments used in the reported ,experiments. (Levy Tr. at423:20-424:12; DTX23 at 655 (under"Experimental Protocol" and subheading · "Emulsified Reactions")) For example, the Tawfik and Griffiths paper provides precise amounts of specific reagents to use, identifies the temperature to use, and details the mixing procedure. (DTX 23. at 655) . · 122. The same paper also .explains tha~ "[t]he precise stirring setup can greatly affect droplet size:" (Levy Tr. at 424:13-19; DTX23 at 655 under "Experimental Protocol" and subheading "Emulsified Reactions")) Apatent by Tciwfik and Griffiths that issued prior to the '071 provisional filing date also explains that "the size of the emulsiOn microcapsules niay be varied simply by tailoring the emulsion conditions used to form the emulsion according to the . requirements of the selection· system~" (DTX 24at11:29-31) From these disclosures, a person . . of ordinary skill would have understood·the size of the emulsions reported in the Tawfik.and · Griffiths paper could have been controlled bymanipulating the emulsion conditions and the 36 ·stirring setup. (Levy Tr. at427:4-17) 123. Ghadessy .et al published a.paper in PNAS - a well read and respectedj ournal - that modified the .emulsions reported in Tawfik and Griffiths. (Id. at427: 18-430:5; DTX 33 .at · 45:53, ·heading "Emulsification of CSR," referring to reference (16), which is identified on page 4557 .as DTX 23) The Ghadessy paper states:· We used the recent1y described water-in-oil .emulsions (16 IDTX 23]) but modified the composition of the surfactants as well as.the water-to-oil ratio. This modification greatly increased the heat stability .and allowed PCR Yields in the emulsion to approach those ofPCR in solution. Compartments "had average diameters of 15 µm a:nd proved heat-stable ....... . (DTX 33 at .4553, below ·"Principles Underlying ·CSR~') 124. By modifying the emulsion of DTX 23, the Ghadessy authors were able to use the .emulsions for PCR amplification. (Levy Tr. at 430:20-431 :5) By slowing down the speed of the stir bar, the authors were also able to _significantly increase the ~ize ofthe emulsions reported in . . . DTX 23. (Id; at 431:6-14, 453:4-17) · The Ghadessy paper demonstrates that size optimization of · emulsions is a straightforward process.- (Id. at 431 :15-20) . 125. . A person of ordinary skill in the art would have been familiar with DTX 23, DTX · 24, and DTX 33 at the time the '071 ·provisional was filed. (Id. .at 432:14-23) The JHU '690 ·application references DTX 23 and DTX33 as examples of how to form emulsions. (ATX 1003 "if 44 (reference 14 is Tawfikand Griffiths (DTX 23) and reference 15 is Ghadessy et al. (DTX 33)) ·Using those references,. a person of ordinary skill could have manipulated the size of the . . . emulsions reported in DTX 23 or. DTX 33 to encapsulate a bead without undue experimentation~ (Levy Tr. at 432:5-13) There is no conceptual difference between encapsulating a bacterium or a 37 bead . .(Id. .at 431:21~432:4) 3. The ·'.071 Provisional J>esc:ribes and Enables 'St~ps (c) .and (d) 126.. · Step (c) oftheCountrequires amplifying the fragment of step (a) in the microreactor. (Id. .at 433:15-20) The "'071 provisional describes step (c)ofthe Countbecause it ·describes emtilsionPCR. (Id. at 433:12-25; ATX 1015 .at 46-47) 12 7.. The '071 provisional describes _step {d) because it describes pifophosphate sequencing. (Levy Tr. at434:1-16; ATX 1015 at38) In order for pyrophosphate sequencingto work, the DNA has to havebeen amplified. (Levy Tr. at 434:6-9). 128. ·The '071 provisional contains no indication that a partial digestion should be performed, as opposed to a complete digestion. (ATX 1015 at 40) . M. The '240 Application :Describes :and Enables the Full Scope ·of the . Count 129. The '240 application describes and enables the full scope of step (a) of the Count . because it describes and enables making genomic DNA libraries. or.cDNA libraries derived from anypopu1ation of nucleic acids.· (Levy Tr. at394:11-395:7; ATX 1001at3:25-28) A person of ordinary skill in the -art would have understood that methods of generating those libraries involve . the formation of more than one copy ofa particular DNA fragment. (Levy Tr. at 395:8-1.5) . 130. The '240 application also incorporates by reference the '071 and '592 provisionals. 7 (Id. .at396:6-15; ATX 1001 at 1:3-6) It also incorpor~tes by reference U.S. Patent No. 7,323,305, which Gontains the entire disclosure of the '"592 provisional. (Levy Tr. at 396:16398:12,400:11-20; ATXlOOl; DTX 12; DTX 13) 7 Given that the '071 and '592 applications describe and enable the preamble, the '240 application does as well because it incorporates these disclosures by reference. 38 131. The '240 .application describes delivery of single~stranded and double-stranded . DNA to the microreactors by describing pre-hybridizing beads via oligonuc1eotide primers attached to the beads. (Levy Tr. .at404:10.:.1s; 7:16-8:31, 25::6~26:30) The '':592 and ~071 provisional applications _also describe .delivery of DNA pre~hybridized to-.a-bead, so those app~ications also describe the delivery of single-stranded and double-stranded DNA to the -microreactor. ,,(ATX 1013 at21:1~10; ATX1015 at 46-47) When theDNAfragmentis pre-hybridized to the bead via that primer, it forms a short region of double-stranded DNA. (Levy Tr. -at 404:16-22) The other portion of the fragment is single-stranded. _(Id. at 404:23-24) Thus, a partially single-stranded and partially double-stranded· DNA fragment is delivered to the microreactor. (Id. at 404:25-405:3) 132. The only difference between delivery of a single-stranded DNA fragment - ' ' pre-hybridized to .a b~ad and.a double-stranded DNA fragme11t-tiot pre-4ybridized is the omission of the-pre-hybridization step. (1Yagi Tr. at 139:5-10; Levy Tr. at405:17-22) The components· are still delivered-the same way. (Levy Tr. at 405:17-22, 406:16~20, 475:10-24) -133. Very little, ifany, experimentation would be required to deliverthe double-stranded DNA, as opposed to the single-strandedDNAfragment. (Levy Tr. at 405:25-406:20, 437:15-21) Dr. Tyagi's speculation that 'delivering double-stranded DNA _separately from a bead would be different from delivering DNA pre-hybridized to .a bead is notbased on any actual experimental evidence. (Tyagi Tr. at 547: l 6~548 :5) 134. Dr. Levy explained that modification- of the parameters necessary to successfully form a microemulsion could be done in a day. (Levy Tr. at 456:4-14) Dr. Levy further opined that modification of those parameters is "extremely simple," and would "absolutely not"_ require 39 undue experimentation. (Id. at :501: 17-502: 1) 135. Dr. Tyagi testified that .delivery of one DNA frel:gment and one bead into each ofa ,plurality ,of microreactors could be confirmed using "statistical probabilities."". (Tyagi 60:12-15) The ''240 application uses statistical probabilities to confirm deljvery of DNA and beads to themicroreactors. (ATX 1001at16:12-17:27) N. .JHU is Not ·Entitled .to .a ,Priority .Date Earlier than .June 5, 2003 136. None of the priority evidence JHUprovides demonstrates that JHU had .conceived of all the elements of the Count prior to June 5, 2003, induding: (1) delivering a plurality of · molecules ofthe ~ame fragment ofDNA "into microreactors; (2) any type of emulsion; (3) any microreactor; (4) _having a single head in a microreactot; and (5) having a single molecule of a DNA fragmentin·a microreactor. . ' ·13 7. .·JHU .cites isolated testimony and documents relating to PCR pre~amplificatiort outside of an emulsion, or optimizati<?n of emulsion conditions,· but :there is no .evidence the JHU · inventors had the concept of putting template into the emulsion, amplifying in emulsion, and detecting amplification products from the emulsion, which are all required elements of the Count. 13 8. ATX 2043 and/or ATX 2081 are not .evidence for performing emulsion PCR, prior to June 5, 2003, as these documents contain no explanation, nor ~s there.any·non-inventor corroborating testimony explaining the experiment underlying the data, or the computer files. 139. Neither the Dressman publication (ATX2026) nor any of its drafts can corroborate work done prior to the date the manuscript was completed, and its existence has only been proven by JHU as of June 5, 2003. 40 140. The oniy .non'.'"inventor testimony JHU has .cited are th~ dedarations of Leslie . . ' . ..... Mesz1er (ATX2029) and.JasonBriody (ATX2081). 141. · Declarant Meszler merely testifies regarding .calendaring time for inventor Dressman to use :a flow.cytometer in the Cell Imaging Core atJHU and the charges for that use, .and regarding a listing of data folders from a.Maclntosh . computer used at the Cell Imaging Core in 2003. (ATX 2029) 142. DedaranfBriody likewise testifi~s merelyto a listing of data folders from the Macintosh .computer and associated metadata. (ATX 2081) 143'. Neither of the corroborating declarations includes any information as to the . . . . . . . · experiments that were being performed ~y inventor Dressman in the Cell Imaging Core, nor .do they address any ,aspect of the invention of the Count. 144. T~e Januazy29, 2003 email from Dr:Vogelsteinto Dr. Kinzler (both inventors) does not refer to various required ·ele~ents of the Count: (1) delivering a pluntlity of molecules of the same fragment of DNA into microreactors; (2) anytype of emulsion; (3) any microreactor; (4)having a single bead fa a microreactor; or (5) having a single molecule of:a DNA fragment in a microreactor. (ATX 2039) Furthermore, Dr. Vogelstein admits that the attachment to the email relates to a non-emulsion format. (A'IX 2034 '~ 12} Moreover, the January 29, 2003 email is not corroboratedbya non-inventor. 145. The February 10, 2003 PCR amplification of the Calpain gene from a heterozygous patient source provides no indication as to the future use of the amplification products, and thus, no evidence was submitted that the Calpain amplification produds were · intended for use in emulsion PCR. Moreover, none ofthe inventor testimony regarding this 41 corroborated. (ATX 2040,.ATX 2034 ifif 27-28, ATX 2051if29; ATX.208o·ir 5)_ .146~ i .JHU' s .contention that Dr. Dressman µsed the amplified °ialpain gene for a bead emulsion ·ex_periment on :February 25, 2003 is based .so1e1y on. uncorroborated inventor testimony. Moreover,thedted pages ofDr. Dressmari's not even refer to using an emulsion, much less to microreactors containing a single DNAfragment.and a single bead .....{ A TX 2051 :if 29; ATX.2036 at39) 147. JHU cites other isolated testimony and documents that relat~ to PCR · pre"".amplification outside of an emulsion, or optimization of emulsfon conditions, as alleged evidence ofconceptimyin February 2003, but there is no- corroborated evidence that the inventors . . hadthe concept ofputting template into the emulsion; amplifying in emulsion, and detecting amplification products from the emulsion, which are all required elements of the Count. 148. There is no corroborated evidence that JHU performed bead emulsion PCR on March 10 and March 31, .2003, using a single bead and a single template DNA in niicroreactors ·formed ina water-in-oil emulsion. (ATX 2024; ATX 2032; ATX 2051ifif26-55,ATX 2079 ··irir 32-39) 149. ·with respect to the March 10 experiments, there is no evidence of: (1) multiple c()pies of a particulartemplate.{i.e.; generated as by PC:R); (2) a single template; or (3) a single bead per compartment, as required bythe Count. (ATX2036 at 71) 150. .· With respect to the ·March 31 experiments, the cited evidence refers orily to a PCR product, not to its source. (Id. at 99) Furthermore, there is·no discussion and/or evidence of a single DNA template molecule and·a single bead in a water'.'"in-oil emulsion mfororeactor, and there is no evidence that the product was detected, as required by the Count. (Id.) . 42 1 151. The last dated page in Dr. Dressmati's notebook is from April 1, 2003. 152. .JHU cannot rely on ATX 2081 as evidence for performing emulsion PCR .. experiments, prior to June 5, 2003, because it is merely a listing of files recovered from a Macintosh computer, suggesting that Dr. Dressman had the .JHU Cell Imaging Core perform some sort of flow cytometery, but there is no evidence in ATX 2081 regarding what was analyzed in any of those.flow cytometry runs. Moreover, there is no corroborating evidence regarding what was analyzed in any of those flow cytometry runs. r 153. JHU cannot locate information allegedly contained in a three-ring binder of Dr. Dressman to explain what experiments were listed in ATX 2081. (ATX 2079 ifif 4-5) 154. Likewise, JHU cannot rely on ATX 2043 as representing data associated with emulsion PCR experiments, because there is· no explanation in the document or in any non-inventor corroborating testimony that explains what type of experiment the data represents. There is no evidence thatit represents bead emulsion PCR or using microreactors containing a single bead and a single template DNA. 155. · JHU cannot rely on ATX 2042 as evidence of another alleged bead emulsion experiment prior to May 12, 2003,, because it is an undated draft of a manuscript that is not corroborated by non-inventor testimony. (ATX 2034 ·irir 31-38; ATX 2080 ·ir 6-7) 156. JHU has not provided evidence that there was an emulsion PCR experiment performed on May 20, 2003, because JHU relies solely on uncorroborated inventor testimony and an undated draft of a manuscript that is not corroborated by non-inventor testimony. (ATX 2034 ifif 31-33; ATX 2051 if 58; ATX 2079.ifif 27, 29; ATX 2080 if 6) 157. The JHU inventors use hindsight to attempt to reconstruct the evidence they could 43 -not.provide, by citing to the Dressman publication (ATX 2026) and proposing dates on which the workreflected in the paper might have occurred. However, neither the Dressman publication (ATX 2026) nor any ofits uncorroborated drafts(ATX 2042 andATX2097through2114) can corroborateworkdone-priorto the date the manuscript was completed~-and its existence has only been proven byJHU as of June 5, 2003. 15 8. JHU has not provided any evidence-that 'it continued to work on bead emulsion after the June 5, 2003 manuscript. ,DISCUSSION - I. _ PRIORITY _ A. :LegaLStandards JHU, as thejunior party to the Interference, must show priority by a preponder<;lllce of the - - - evidence. 8 See Brown v. Barbacid, 276F.3d1327, 1332 (Fed. Cir. 2002) ("Brown I"). -"Priority of invention .and its constituent issues of conception.and reduction to practice are questions of law predicated on subsidiary factualfindings." Singh v. Brake, 3 l 7 _F.3d 1334, 1340 (Fed. Cir. 2003). "Priority of invention.' goes to the first party to reduce an invention to practice unless the _ otherparty can show that it was the first to [ 1] conceive the invention and [2] that it exercised reasonable diligence in later reducingthaflnventionto practice:'" Brown 1,276 F.3d at 1337 (quotingPrice-v. Symsek, 988 F.2d 1187, 1190 (Fed. Cir. 1993)); see also Brown v. Barbacid, 436 F.3d.1376, 1378 (Fed. Cir. 2006) ("Brown II") ("The party that is first to conceive the 8 An applicant who challenges an issued U.S. patent during an interference must prove priority by clear and convincing evidence. See Bruning v. Hirose,-161F.3d681, 684 (Fed. Cir. 1998). -The patents in this case were co-pending and had not yet issued at the time the Interference was initiated, so the preponderance otthe evidence.standard governs. See id. at 686. 44 - invention in interference, if las.t to reduce the invention to practice, is entitled "to .the patent based . on prior conception jf, as· first to. conceive, ·he :exerCised reasonable ·dili.gence from 'atime· bt'.fore . . the .other party'.s .conception .date·to his .own reduction to :practice date:"). ·"Conce.ption is the formation, fa the mind .of the inventor, ofa defiriite and _permanent . . . idea ofthe complete and operative invention, ,as it js thereafter to be .applied in practice." In re Steed, 802 F.3d 1311, 1320 (Fed. Cir. 2015) (internal quotation marks omitted). To show conception, ':a -partymustshow:possession of.every feature recited in the.count, and that.every · limitation of the count must.have been knowp. to the inventor at the time of the alleged conception." Coleman y'. Dines, 754 F.2d 353., 359 (Fed. Cir. 1985) . . When proving conception, the inventor·"must provide independent corroborating evidence in addition to 'his own statements and documents~" Martek Biosciences Corp. v. . . . Nutrinova, Inc., 579 F.3d 1363, 1375 (Fed. Cir. 2009) (internal citation omitted). Corroboration . is subject to a·"rule.ofreasori'' analysis. Reese v. Hurst, ·661F.2d1222 (C.C.P.A. 1981). "Accon;:lingly, .a tribunal must make a reasonable analysis of all of the pertinent evidence to . . . determine whether the inventor's testimony is credible:" Kridl v. McCormick, 105 F .3d 1446, 1450 (Fed. Cir. 1997). 2. Reduction fo ·Practice "A reduction to practice can be either a constructive reduction to practice, which occurs when a patent application is filed, or an actual reduction to practice." Cooper v. Goldfarb, 154 F.3d 1321, 1327 (Fed. Cir. 1998). "In order to establish an actual reduction to practice, the inventor must prove that: (1) he constructed an embodiment or-performed a process that met all . 45 . . . · the 1imltat~ons of the interferen9e count; and (2) ·he determined that the inventio~ wo~ld work for .its intendedprirpoSe ..,' Id. When provin.g a . constructive reduction fo practice, "all that is necessary for a party to be . . entit1edto benefit of an earlier filed application for priority purposes is .compliance with35 U;S.C. § 112 ·With respect to at least one embodiment within the scope of the count." Palko-Gunter Falkner v. lnglis, 448F3d1357., 1362 (Fed. Cir. 2006) (internal brackets and quotation marks omitted). h1Huntv. Treppschuh, 523F:2d1386, 1389 (C.C.P.A. 1975), the Court of Customs and Patent Appeals distitl:guished 'between -requirements under § 112 for applications ser"Ving as bases for priority, on the one hand, .and interfering .applications which · have the potential to ripen into -issued patents,· on the other: Hunt's :parent :application is relied upon as a prior constructive reduction to practice; whereas in_ [Smith v. Horne, 450 F.2d 1401 (C.C.P.A. 1971)], the disclosure was relied upon for a right to make the count. In the latter situation the requirements. of . the first paragraph of 35 U.S. C. § 112 must be satisfied for the full scope of the .count. Inthe former, however, the § 112, first paragraph requirements need only be met for an embodiment within the count. · As applied to this case, the holdings in Falko and ·Hunt require 454's '592 and ''071 applications _,.which 454 relies on only as bases for priority-'·to describe and enable at least one embodiment . . . ·· within the scope of the Count. By contrast, 454' s '240 application must describe and enable the · full scope ofthe Count, as 454 desires· to obtain· (in fact, retain) patent rights to the claims set out in the '240 application (which is now the issued '305 patent) .. 46 ·B. · 454'-s1.Conception and Reduction to Practice l. 4541Conceived .of ~he Inve~tion 'by December 20, 2002 Considering the evidence in its entirety, the Court.concludes that454 ·has proven by·a ·preponderanc.e ofthe evidence that it had a "definite and permanent idea of the complete and operative :invention,'' Steed, 802 F.3d .at 1320, by no later than December 20, 2002. (See FF3555) 9 A group of the inventors initiall.y discussed the idea for a method for analyzing nucleic aCid sequences using bead emulsionPCR on June 7, 2002, as evidenced in Dr. Berka's lab notebook andastestified·to byDrs. Sarkis.and Leamon. (ATX 16; Sarkis Tr. atJ80:8-183:5; ATX ll 14'~ 17) Dr. Bef.ka'slab notebook includes a depiction of the basic concept of the invention, . . showing a·PCRreaction involving .individual bead-fragment combinations in a water-in-oil emulsion.· (ATX1094 at 16) Initial experiments performed by Dr. 'Leamon revealed problems with emulsions "crashing" before amplification or sequencing could be performed. (FF39) However, later experiments improved the stabilization of the emulsions, permitting Dr. Sarkis to. complete steps· (a) through (c) ofthe Count in an experiment on December 20, 2002. (FF39, 41-44) Dr. Sarkis used test fragments that had been prepared by Mr. Altman, whote_stified that the test fragments he had had been PCR amplified and, therefore, comprised multiple copies of the same DNA fragment, in compliance with step (a)~ (FF44-48) Dr. Sarkis testified that.the goal ofthe454 . inventors was clonal amplification, meaning their goal was to have a single eff<?ctive copy per . . . bead per microreactor,_ in compliance with step (b ). (FF54) · Dr. ·Sarkis and Mr. Altman had 9 Citations to the Court's Findings of Fact, which are provided·earlier in this Opinion, are in the following format: ·"FF[paragraph number(s)]Y 47 previously sequenced the test fragments in separate experiments, in compliance with step (d). Although sequencing was not completed on December 20, 2002 on the amplified copies of DNA, the ultimate goal of the 454 scientists was to develop .a procedure that would include step (d) of the Count (and, again, they had done step (d) of the Count earlier and, plai~ly, still had · possession of step {d)). (See Sarkis Tr. at 201:1-11) Dr. Sarkis.'s testimo11y, which the Court found to be credible, in combination ·with all of the other relevant evidence, indicates that 454 conceived of the invention by no later than December 20, 2002. · 454's conception was corroborated by non-inventor testimony. (See FF56-87) In particular, Mr. McDade corroborated the discussions of.June 7., 2002. (FF77) In addition, Mr. McDade specifically recalled that Dr. Sarkis performed a bead emulsion PCR experiment that resulted in template amplification and sequenceable product just before Christmas in December 2002. (FF78) Ms. Lanza recorded in her lab notebook the sequence of_ one of the PCR-generated test fragments used by Dr. Sarkis in his December 20, 2002 experiment. (FF47) Mr. Altman testified that he PCR .amplified the test fragments used by Dr. Sarkis in his December 20, 2002 experiment. (FF46) 454's conception is further corroborated by testimony regarding the applicability of the Poisson distribution to Dr. Sarkis's December 20, 2002 experiment. (See FF56-60) The Poisson distribution may be relied on as evidence of what actually transpired during the experiments performed by454's scientists, including the December 20, 2002 experiment. (Levy Tr. at 361:16-22, 498:13-24) The Court credits Dr. Levy's testimony and finds his discussion of the Poisson distribution to establish that it is highly probable that Dr. Sarkis's December 20, 2002 experiment resulted in more than one microreactor containing a bead and the same DNA 48 fragment, _satisfying the :limitations of step (b )- of the Count. JHU'_s . .criticisms of 4~4 's experimental evidence from the December 20, 2002.,experiment are contradicted -by documentary evidence and testinl.o~y from-454' s witnesses, .all of-whom ·the. Court found to be-credible. (See, .e.g.~ D.I.110 at.31) (alleging there is '~[ri]o evidence of .amplification oftest fragments .according to step (c)," when evidence shows that opposite is true (seeATX 1105.at95-96)) Regarding conception specifically, JHU alleges that the 454 inventors failed to appreciate step (a) of the Count and, therefore, did not.conceive of the invention ~y December 20,2002. (See D.I. 110 at28-32) The Court disagrees. Dr. S-arkislrnewthetest .fragments were PCR-:generated and_ included them in _his microemulsion experiment, according to :step·(a) ofthe,Count. -(Sarkis Tr. at 194:11,-15) 10 This s.atisfies step (a) of-tbe Count as it :plainly involved generating a plura1i~y of molecules of a fragment of DNA. ·JHU focus~s mi the fact that 454 was ultimately interested "in analyzing adenovirus fragments that were not .generated _in such a way that would have created more than one of the ·same fragment accorditigto step (a). (See id. at 28-29) Regardless of what the ultimate goal of 454 was, as a :company, the evi_dence shows that Dr. -Sarkis understood himself to be performing · step (a) ofthe Count ~y including test fragments in his microemu1sion experiment on December 20, 2002. Step (a) was known by, .and in the possession of, Dr. Sarkis as of that date. JHU's 10 JHU"insists that both "conception and reductionto practice 'require contemporaneous recognition and appreciation of the limitations of the claimed invention' not just fortuitous inheren~y." (D.I. 115at29)(quoting Mycogen Plant Sciences; Inc. v. Monsanto Co., 252 F.3d 1306, 1314 (Fed. Cir. 2001)) However, as 454 argues, the Federal Circuit-citing Mycogen has held thafin an interference a prior inventor need not "demonstrate that [she] recognized the exact language of the ultimate count" but "onlythe subject matter of the invention:" (D.I. 118 at 27) (citing Henkel Corp. v. P&G, 485 F.3d 1370, 1375 (Fed. Cir. ~007)) 49 insi~terice that it and 454 had different aims .and were pursuing different invenfions 11 does . . nothing to undemiine the Court's findings :that 454 :conceived of±he invention of the Connt l~y 'December 20, 2012 12 -,even Jfat some earlier date .th~parties'' ·initial.intentions ,d_ijfered. . JHU raises :additiona1 challenges to -4:54' s conception ofthe invention which .are addressed .below in .connection With the court's analysis of 454' s reductions ·to ·practice. 2. _454 Actually Reduced the lnvention to Practice by .January 15, 2003 454 ·has proven ~ya preponderance of the evidence that it actually reduced the-invention . ofthe Cotintto practice, starting at least by.January 15, 2003. _(See FF61-87) For example, from .J anuazy 14 to 15, 2003-, Dr..Sarkis set up .emulsion PCR reactions using a combination of . . . . . . - . adenovirus DNA and test fragments (TF4, TF6, TF7., and F6) with an input of600,000 beads and 1.2 million molecules of DNA {two copies per bead) and successfully obtained sequencing . . . results for th~ F6 test fragment.from emulsion-PCR-generated beads. (FF62) Th_e F6 fragments · AccordingtoJHU,·"[t]he objective ofthe JHU application is to·findmuta~ions in· spec:ific portions of the genome," which "requires:generatingmultiple copies of a fragment of the · genome' through two steps: "pre-amplification," which is step (a) ofthe Count, and "emulsion amplification," step (c). (D.I. 110 at.1) .JHU continues by.contendingthat the objective . of 454's application, by contrast, "is to assemble genome sequences from snippets of the genome;" which ~entails "the genome be[ingJ fragmented into many overlapping pieces." (Id.) In ffiU' s telling, the "use. of a second or 'pre-amplification' step· is counter...;productive to the objective ofthe 454 application as it generates multiple copies of the same fragment," whereas the whole point of 454's invention is to ·"piece[] together" multiple fragments "to assemble the .entire genome." (Id.~; see also D .I. 115 at 18 ·(" [JHU] has produced substantial evidence at triai that not only are the methods described inthe [454] '071 _and '592Applications different from ffiU' s applications, but that the reason for that difference is due to the different goals and . objectives of the applications-gene sequencingv. detection of gene mutations-which shows the methods are not obvious variants of one another.")) · 11 ' . . . 12 - ' ' ' . This is months before JHU's·conception-which occurred no earlier than Jurie 5,··2003....:. meaning that 454 both conceived the invention of the Count before JHU did and then began using·reasonable diligence to reduce to practice before JHU even con.ceived of the invention. See· Brown II, 436 F.3d at 1378. 50 satisfy step {a) ofthe Count, because the.y were ,generated by PCR, .as Mr. Altman explained. (FF46) Accordmg:to Dr. Levy':s testimo11y, which the :Court credits, this experiment would'have -produced .at1east two microreactors (i.e., a pluratity) eacb containing one bead and one of the same F6 test fragments, in accordance with step (b) of the Count. ·(Leyy Tr. at 359:23-360:11) The F6 fragments, bound to beads; were then .amplified .and sequenced in accordance with _steps (c) .and (d) of the Count. ·(ATX 1105at112.,.13) Later experiments performed under similar .conditions were additional, subsequent · reductions to practice. (SeeFF63 (January 23, 2003 experiment); FF64 (January28-31); FF65 . . . - . ' . (February5); :FF66 (February 6}; FF67 (February 7);EF72(February19); FF73 (February27); FF74 (March 7);-FF75 (Marcb28)) .JHU 9ritidzes 454's evidence ofconception and reduction to .practice in.a variety of ways. (See, e.g., D.I. 110at 30.:.31) JHU argues thatthe454 experimental documents do not show a conception or reduction to practice of step (b) ~'based on work done with the test fragments." (Id . .at.30). The Court disagrees. As Dr. Levy opined,thetest-fraginents were included in - · microemulsions as required in step (b) of the Count. (Levy Tr~ at 359:23-360:11; see also Sarkis Tr. at 194:11-15) ·- . JHU contends that the "only ·basis that Dr. Levy ,and 454 provide" to show conception and · reduction to practice of step (b) is thatthe·Poisson distribution theory:applies to the distribution . of fragments in the microemulsion. (DJ. 110 at 30) Even if this allegaticm were ~e (it,is not), .JHU' s own expert testified that the Poisson distribution is "alwaystrue~' (Tyagi Tr. at 13 7 :2-3, 138:25-139:1), and the Court credits Dr. Lery's testimony that the Poisson distribution would ·_ hold true with respect to 454' s experiments that res_-u.lted in reduC-tions to practice of the invention 51 ·(Leyy Tr. ~at 359:23-3 60 :11). JHU .argues that "the .data.collected {number of fragments per bead)" during the 454 .experiments ~"is insufficient to .show that the Poisson distribution was actual~y achieved." (D.I. 110 at 30) Again, the Court credits Dr. Ley)'' s testimony that the Poisson distribution would hold true.and predicts two or more niicroteactors, compJying withthe.reqriirements of step (b). ·(Levy Tr. at359:17-22) As explained ~y Dr. Levy, .one .could .estimate the number-of .compartments in .an emulsion b.y "knowing the diameter of those compartmen~s . and the volume . of the aqueous solution going in~" (Jd.. at365:ll-22) Based on the number of compartments, the number offr<l:gments, .and the number of beads, Dr. 'Levy was able to .conclude that the 454 reductions to-practice would have included more than one compartment with a sing1e head and the same fragment of DNA, "because you're able to do PCR that's detectable in the sequencing .action:" (Id ..at365:23-366:15) The Court further credits Dr. Lery's opinionthatthe very factthat45_4 obtained sequencing results necessarily. presupposes that some nonzero number of identical test fragments were delivered to microreactors'. As Dr. Levy explained, while 454' s experiments ·were not · designed totestthe Poisson distribution, .and therefore cannot demonstrate the percent of . instances. in which there is a single fragment on a single bead, the experiments do show ·"sequence data." (Levy Tr. at 360:9-361 :3) This is itself confirmation of there being at lea~tone . . . . . . . . . instance ofasingle fragment ·on a sillgle bead:.''.'[I]t is very clear that you only get sequence data · from the experimental setup. You only get sequence data if there is one fragment on one ·bead going into the .amplification." (Levy Tr. at 360:20-24; see also id. at-496:8~12 ("You can't use this data to back-calculate the percentages, but it absolutely assliredlytells you thatthere were 52 individual beads with individual fragments thatamplified and were capable ofbeing sequenced and could be mapped to the, in this case, the identifier genome.") Dr. Tyagi ultimately agreed that if one ,gets a sequence (as 454 did), this means that there was only ,one sequence on the bead. (1)agi Tr. at 546: 6-8 ("I would agree with Dr. Levy, that sequence, if you get a sequence, that means there was only one sequence on the bead.")) JHU argues that the fact that test fragments were added to emulsions separately from adenovirus means amplification and sequencing would have produced "exactly the same signal" whether there were 1 or 100 fragments in a microreactor. (D.I. i10 at 31; Tr. at 577:3.:.10) JHU's.arguments on this issue are vague but appear to reflect its broader speculation that there is no way to-be sure step (b) was satisfied by two or more microreactors containing only one of the . ' same DNA fragment anda single bead. The Court credits the testimony of Dr. Sarkis, as well·as the documentary evidence from Dr. Leamon's notebook, and is persuaded the record demonstrates compartments of"about the right size" for containing single bead-single fragment combinations. (Sarkis Tr. at 190:5-191:10; see also id. at 197:2-7) Moreover, the Court credits Dr. Sarkis's testimony that his January 14-15 experiment produced at least two microreactors complying with step (b). (Id. at200:15-22) JHU asserts (without showing) that there is no evidence that the beads to which the test fragments were allegedly attached were capable of hybridizing to the test fragments. (D.I. 110 at 31) The Court disagrees. Dr.· Sarkis's lab notebook and testimony from Dr. Berka indicate that 454' s standard practice was to wash and perform other actions meant to remove unbound beads, leaving only the beads and fragments bound to them (if any) for analysis. (See generally ATX 1105 at 112; Tr. at 306:23-313:12 (noting that, after washing and PCR cycling, beads and any 53 . . fragments bound thereto were analyzed, not unbound fragments); see also Sarkis Tr. at 217:1-4) JHU further contends in its post-trial.briefing (without citing to the record) that the sequencing primers used for test fragments raise questions about whether the 454 experiments practiced the Count. (D .I. 110 at 31) ·454 responds that this argument was never presented at any earlier point ·in this case, and that there is no record evidence cited in support of it, so JHU' s position in this regard is not. entitled to .any evidentiary weight. (D .I. 118 at 28) The Court agrees and will disregard JHU' s attorney argument - which is unsupported by anything in the record. 3. 454'-s ·conception and Reductions to Practice are Corroborated 454 has shown by a preponderance of.the evidence that it conceived and reduced to practice the invention of the Count as described above and has come forward with credible, persuasive non-inventor evidence to corroborate the experiments discussed above. (See FF7687) Mr. A1tman corroborated_production of.the F6 test fragments by PCR, satisfying step (a) of the Count. (FF46) Ms. Lanza recorded the sequence of the F6 test fragment in her lab notebook, evidencing conception of step (d) and the intention of the experiments to amplify and detect F6 test fragments. (FF47) Dr. de Winter corroborated all steps ofthe Count with his testimony that his team performed emulsion experiments either with test fragments alone or with test fragments m!xed with adenovirus. (FF53) Dr. Lanza corroborated reductions to practice in January 2003 a~d testified that she worked. "very closely" with Dr. Sarkis at the time. (ATX 1126 ifif 10-13) Mr. McDade corroborated conception as of December 20, 2002, as already discussed above, and the subsequent reductions to practice. 54 iC. · 454's Priority 'Claim io '~·071 Application ·454 has proven that itis entitled to claim .priority to the ., 071 application, because it has -shown thatthe '071 application describes and .enables a person of ordinary skill in the art to practice at least one embodiment within the .scope ofthe'Count. (FFl 07~28) The "'071 .application,discloses use.of restriction endonuc1ease_digestion for the creation oftemplateDNA used in conformance with step (a) of the Count. (FF109, Tl4) The ~071 also disdoses use of .commercial~y available -kits for DNA isolation, which would.have necessarily induded kits that havemorethan one of the· same DNA fragment.· (FFlll) Use of restriction enzymes on DNA fragments from commercially available kits would inpracticing step (a) of-the Count. . (FFll 4-15) The ,, 071 application discloses step (b) of the Count. (FF11 7-25) The '071 . appli~ation . specifically states: ~'The.average size ofthe emulsion capsules must be optimized to maximize . . the number of single beads containing _single strands of DNA, that c;an:be incorporated within a single emulsion vo1ume." (FFl 17) The foregoing refers to use of the DNA template generated . ·during step (a), as described above.· (FFl18) The-'071 application discloses enough guidance "and the :necessary parameters to ~nable someone of ordinary skill in the artto practicethe Count. (See FFl 19-25) . Finally, the '071·application discloses steps (c) and (d) of the Count, as described in the offact. (FF 126-2 7) JHU argues that the '071 application does not disclose the full scope of the invention . because it does not disclos~ methods irivolving double-stranded DNA template used in step·(a) or use of non-pre-hybridized DNA separate from beads delivered to microreactors according to step 55. (b). (D .I. 110 at 10) JHU' s contentions rely on a flawed legal premise. In order to support a claim of priority, it is not necessary for 454 to prove written description and enablement of the full scope of the Count qythe '071 application. See Hunt, 523 F.2d at 1389. Instead, 454 need only prove written description and enablement of an embodiment - which 454 ·has done, as explained .elsewhere in this Opinion. Nevertheless, the Court finds that the '071 application's disclosures do enable even what JHU characterizes as the "full scope" ofthe invention. Regarding.JHU's· arguments as to step (a), the -'071 application discloses use of single. stranded DNA hybridized to a bead via oligonucleotide primer such that a short region ofdoublestranded DNA would be created at the bond. (Levy Tr. at404:10-22) Thus, the '071 application discloses both double- and single-stranded DNA. Moreover, Dr. Levy testified that delivery of only double-stranded DNA to an emulsion instead of single-stranded DNA would not have any effect on the outcome of the method in the Count. (Id. at 405:8-16) Therewould be no significant difference in how one would perform the method of the Count by using only double- . stranded DNA in step (a). (Id. at 405:17-406:3) With respect to JBU' s argument that the '071 application does not describe or enable · delivery of DNA separate from a bead into a microreactor, the Court is persuaded by Dr. Levy's . . opinion that a person of ordinary skill in the art would know how to deliver DNA separate from beads, given the disclosure of a pre-hybridized embodiment, such as the one in the '071 application. (Id. at 406A-20) JHU does not argue that the '071 application (or the '592 application) fails to describe or enable steps (c) or (d) of the Count. 56 · D. 454.'s :pf:forify 'Claim to '592 Application 454 has _proven that it is entitled to claim prioricy to the ·'592 application, because jt has shown:that the ''.592 application. describes .and enables a person of ordinary skill in1he art to practice at least one enibodimentwithlnthe scope of the Count. -.(FF107-28) The ·'592 application discloses everything that is in the ., 071 application - plus additional detail. .(Compare ATX 1013 ('592application) with ATX 1015 ('071 application)) Thus, for atleastthereasons given above with respect to the '071 ·application, the '592 application also qualifies as a constructive reduction to practice. E. . JHU'·s Conception :and Reduction to Practice . . The Court. previously determined that JHU conceived of the 'invention on June 5, 2003 -and exercisedreasomible diligence until:reductionto practiee onJuly5, 2003. (D.L 97 at 12~ 16) . JHffhas failed to prove that it is entitled to.apriority date .earlier thaI1June5, 2003~ The only non'""inventor corroborating ~vidence cited by JHU in support of its arguments for an earlier · prionty date fails to disclose the following required elements of the Count, as explained ~y 454 in its answering brief and in the Court's findings· of fact:. ( 1) delivering a pluralify of molecules of . the same fragment of DNA into microreactors; (2) any type of emulsion;. (3) any microreactor; (4) having·a single bead in a microreactor; and (5) having a single molecule of a DNA fragment in a microreactor. (See D.I. '118 at 32;·see also FF140-62) Because JHU has cited insufficient . corrol>0ration of conception and reduction to practice, JHU-has failed tO carry its burden of . proving, by a preponderance of the evidence, an eatlierpriority date. See Marteki, 579 F3d at 1375. 57 iF. . :Conclusion as to .:Priority . . . 454 fois proven.(1) conception by .at least December .2002; (2) actual reduction to practice }Jy .at least January 15, 2003; :and :(3} constructive reductions to practice of the Count ::Qy April 23., ·2003.,·the tiling date of the '071 application, and June 6, 2003,·the filing date-ofthe ''592 application~ As ·previously determined bythe -Court, JHU has proven :a conception .date o{June .5, 2003 and :a reduction to practice date ofJrily.5~ 2003., with diligence .during the interveriing time period suffiCient to support a June .5., 2003 priority date. (D :I. 97 .at 12, 16) Accordingly, . the Court is compelled to hold that454 prevaikonthe parties·' _priority dispute, as454 has proven that it was the first to conceive of the invention ·and the first to reduce it to practice. II. VALIDITY A. Legal Standards l. 'W-ritten Description Paragraph 1 of 35 U.S.C .. § 112 .states in pertinent part: The specification shall contain a written .description of the . invention and of the manner process of making and using it~ in such full, clear, concise .and exact terms as to enable any person . ·skilled in the art to which it pertains, or with which it is most · .nearly .connected, to make and use the same ... :· . [13 ] .and The statute Sets out separate requirements for written description and enablement. ·see Ariad. Pharm., Inc. v. Eli Lilly & Co., 598 F.3d 1336, 1344 (Fed. Cir. 2010) (holdingthat·written description and enablement requirements are separate).. Yet these requirements "often rise and 13 The patent statute was amended in September 2011 by the America Invents Act ("AIA"), Pub. L. No. ·112.;.29, 125 Stat.284, 300-01 (2011). The pre~AIA version of§ 112 · applies in this case._ The post-AIA version of this portion of the statute(§ "112(a)) is identical to the pre-AIA verison. 58 fall togeth~r.~' .Id. at 1352. The parties agree that, in .a case brought under § 146 .arising from an interference, ·invalidityunder .§ 112 mustbe proved ~ya preponderai;ice of.the _evidence. (See Tr. at 31-32, 45) The·Court :~grees that .a.preponderance -of the evidence standard '1PPlies. .See Bruning v. Hirose, 1998 WL 690851, .at *3-5 (Fed. ·Cir. Sept. 29, 1998). Whether a specification satisfies the written .description requirement is .a question offact. See GlaxoSmithKline LLC v.. Banner Pharmacaps, Inc .., 744 F.3d 725, 729 (Fed~ ,Cir. 2014); see also Alcon, Inc. ·v. Teva Pharms. USA, Inc., 664 F. ·Supp. 2d 443, 468 (D. Del. 2009) ("Satisfaction of the written description requirement is a fact..:based:inqriicy, depending on 'the nature of the claimed invention and the knowledge of one skilled in the art .at the time '.an invention is made and a patent .application is filed."') (quoting Carnegie Mellon Un.iv. v. Hoffmann-LaRoche Inc.~ 541 F.3d "1115, 1122 (Fed ..Cir. 2008)). To comply with-the written description requirement, a patent's specification ·"must clearly allow persons of ordinary skill in ihe art to recogriize that the inventor invented what is Claimed." Ariad, 598 F.3d .at 1351 (internal brackets .and quotatio)J_ marks omitted). "[T]he test for·. . . . .sufficien~y is whetherthe disclosure of the application relied upon reasonably conveys to those_· skilled in the art that fhe inventor ·had_ possession of the claimed subject matter as of the filing date..... [T]he test requires an objective inquiry into the four comers of the specification from the perspective of.a person of ordinary skill in the ~rt." Id. "[T]he written description requirement does not demand either examples or .an actual reduction to practice; a constructive reduction to practice that in a definite way identifies the claimed invention can satisfy the Written description requirement." Id. at 1352. However, "a descriptionthatmerelyrenders the invention obvious does not satisfy the requirement." Id. 59 2. Enablenient "Enablement is a question oflaw ~based on underzying factual findings:" MagSil Corp. v. Hitachi-Glob ..Storage Techs., Inc .., 687 F..3d 1377, 1380 .(Fed. -Cir. 2012).· ·"'To he .enabling, the specification of a patent .must teach those skilled in the art how to make ~and use the fu11 scope of the .claimed invention Without undue .experimentation_~.,,,- Id~ (quoting Genentech, Inc. v .. Novo Nordisk, AIS, 108.F~3d1361, 1365 (Fed ..Cir. 1997)) (internal.quotation marks omitted). "Enablement serves the dual function in the patent ~ystem of .ensuring .adequate- diselosure ofthe claimed invention.and of preventing claims broaderthan the disclosed invention.:" Id. at 13 80-81. ·"Thu~, a patentee chooses broad claim language at the peril of losing ar~y claim that cannot-be enabled across its full scope of coverage."" Id. .at 1381. ·"The ·scope ofthe claims must be less .than or .equal to the scope ·ofthe enahlement to ensure that the public knowledge is . enriched by the patent a degree .at least commensurate with the scope of the claims." Id. (internal quotation marks omitted). ''Whether undue experimentation is needed is not a single, simple factual determination, but rather is a conclusion reached by weighing many factual considerations." In re Wands, 858 F.2d 731, 737 (Fed. Cir.1988). These factors include: "(1) the quantity of experimentation necessary, (2) the amount of direction or guidance presented, (3) the presence or absence of working examples, (4) the nature of the invention, (5) the state of the prior art, (6). the relative skill o~ those in the art, (7) the predictability or ilnpredictability of the art, and (8) ·the.breadth of the claims." 1d. Although "a specification need not disclose what is·well knownfo the art;" "[t]ossing out the mere germ of an idea does not constitute enabling disclosure." Genentech, 108 F .3d at 1366. A patent "cannot simply rely on the knowledge of a person of ordinary skill to · 60 serve as a substitute for the missing information in the specification:" ALZA Corp. v. Andrx Pharm., LLC, ·603 E3d 935, 941 (Fed. Cir. 2010) . .3. Anticipation "A patent is invalid for anticipation if a single prior art reference discloses each and every limitation ofthe claimed invention.-" Schering Corp. v. Geneva Pharm., Inc., 339 F.3d 1373, 1377 (Fed. Cir. 2003); see also In re Donohue, 766 F.2d 531, .534: (Fed. Cir. 1985) (stating patent is invalid for anticipation where "each limitation of a claim [can] be found in a single reference, practice, or device"). A "printed ·publication in this or a foreign country" is anticipatory prior art ifit discloses the claimed subject matter "before the invention thereof by the applicant for patent." 35 U.S.C. § 102(a). 14 "The dispositive question regarding anticipation is whether one skilled inthe art would reasonably understand or infer from the reference's teaching that every claim limitation was disclosed in _that single reference." Akamai Techs., Inc. v. Cable & Wireless Internet Servs., Inc., 344 F.3d 1186, 1192 (Fed. Cir. 2003). Whether a claim is anticipated is a question of fact. See Eli Lilly & Co. v. Zenith Goldfine Pharm., Inc., 471 F.3d 1369, 1375 (Fed. Cir. 2006). In this case, which arises from an interference, anticipation must be proved by a preponderance of the evidence. See Bruning v. Hirose, 161F.3d681, 685-86 (Fed. Cir. 1998) ("During an interference involving a patent issued from an application that was co-pending with the interfering_ application, the appropriate standard of prooffor validity challenges is the preponderance ofthe evidence standard."). 14 The Court again refers to the pre-AJA version of the patent st_atute, which governs in this case. 61 "A showing that. a patent was conceived at an- earlier date [than the date of a prior art reference] .and reduced to practice With reasonable diligence is called an effort to swear behind" .a prior art reference. Inc. v. Endicia, Inc., 437 F. App'x 897~ 907 (Fed. Cir. 2011 ); see also Moll v. Northern Telecom, Inc .. , 1995 WL 676420, .at *4 n.1 (E.D. Pa. Nov. 8, 1995) {"[B]y pemiitting.apatent applicant to 'swear behind' the prior reference under 37 C.F.R. § 1.131., .a patent examiner effectively removes the prior a bar to patentability."). "The requirement for the corroboration ofinventortestimony applies to efforts to swear behind a prior art reference:" Stamps, 43TF. App'x .at 908. B. ·whether the '240 Application Meets the Written Description and Enablement Requirements The Court has already determined that the '071 and '592 applications disclose the full scope of the invention, for reasons explained above. Specifically, the Court concluded that the '071 application's disclosure of hybridized DNA to beads discloses both single- and doublestranded DNA and that the -'071 applieation's disclosure of the pre-hybridized embodiment provides written description and enablement support for a non_;pre-hybridized embodiment. (Levy Tr. at404:10-22, 405:8-406:20) The '240 application incorporates the '071 and '592 applications by reference. (ATX 1001 at 3-8) Thus, the '240 application also adequately describes and enables the full scope of the invention.· See Harari v. Lee, 656 F.3d 1331, 1338 _(Fed. Cir. 2011) (holding that patent application incorporated by reference could supply written description support). Thus, while JHU-is correct that 454 must satisfy§ 112's written descriptio!l and enablement requirements for the full scope of the claims it seeks to retain (i.e., the claims of the 62 ·'305 patent, which is the issued·patent that isthe progeny of the '240 application), see Falko~Gunter, 448 F.. 3d at 1362, 454 has met this burden ~by the requisite preponderance .of :the evidence. ,.c. ·whether '240 Application is Anticipated .by the Dressman Reference JHU argues that the '240 applicati~n is antiCipated by the Dressman reference. (D.I. 110 · at l-2) The Court disagrees. The date of the Dressman reference is July 22, 2003. (Id.) The Court.has determined, as discussed above, that 454 is entitled to claim priority to the '071 and '592 .applications for written description and enablement of the full scope of the ·Count. The priority dates of these applications antedate the Dressman reference. Therefore, the Court agrees with 454 that the Dressman-reference is not prior artto .the '240. application and cannot anticipate under§ 102. (See D.I.118 at 7-10) 15 n. ··Conclusion :as to Validity For the foregoingreasons, the Court concludes that JHU has failed to prove that the '240 application is invalid under either of§§ 112or102. ·:CONCLUSION Plaintiffs prove by a preponderance of the evidence that they are entitled to . . ' •' ·. . .. priority. Plaintiffs have also-failed to prove by a_preponderance of the evidence that the '240 application is invalid. An .appropriate Order will be entered. 15 Forthis-reason, JHU's;reliance on Chiron Corp. v. Genentech, Inc., 363F.3d1247 (Fed .. Cir. 2004), is unavailing. Moreover, Chiron is distinguishable, as argued by 454 (D.I. 118 at 8-9), because the separate delivery of beads and fragments o'f either· single- or double-stranded .DNA irito microreactors was not unpredictable, nascent technologylike the technology at issue in Chiron. 63

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