1111111111111111111111111111tM111Q1111111111111111111111111
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`12 United States Patent
`Mathies et al
`
`10 Patent No
`45 Date of Patent
`
`US 7799553 B2
`Sep 21 2010
`
`FOREIGN PATENT DOCUMENTS
`
`EP
`
`0459241
`
`111
`
`121991
`
`Continued
`
`OTHER PUB CATIONS
`
`54 MICROFABRICATED INTEGRATED DNA
`ANALYSIS SYSTEM
`75 inventors Richard A Mathies Moraga CA US
`Robert Blazej Berkeley CA US
`Chung Liu Albany CA US Palani
`Kumaresan Berkeley CA US
`Stephanie H I Yeung Berkeley CA
`US
`
`73 Assignee
`
`The Regents of the University of
`California Oakland CA US
`
`Notice
`
`Subject
`
`to any disclaimer the term of this
`is extended or adjusted under 35
`USC 154b by 0 days
`
`patent
`
`21 Appl No 11139018
`
`22 Filed
`
`May 25 2005
`
`65
`
`Prior Publication Data
`
`US 20050287572 Al
`
`Dec 29 2005
`
`Related US Application Data
`60 Provisional application No 60576102 filed on Jun
`12004
`
`51 Int Cl
`Cl2M1A90
`CUM 1134
`C12M 300
`52 US Cl
`
`200601
`200601
`200601
`4352831 204451 422681
`435912 4352872 4352886 536254
`58 Field of Classification Search
`204451
`422681 435912 2831 2872 2886
`536254
`See application file for complete search history
`
`56
`
`References Cited
`US PATENT DOCUMENTS
`3352643 A
`111967 Ando et al
`
`Soper SA DC WilliamsY XLE SJ assner Y Mang SM Ford
`and RC Bruck Sanger DNA sequencing
`reactions performed in a
`solid phase
`nanoreactor
`coupled
`electnophoresis Anal Chem 1998 70 p 40364043
`
`directly
`
`to
`
`capillary
`
`gel
`
`Continued
`
`Primary ExaminerRobert T Crow
`Assistant ExaminerNarayan K Bbat
`74 Attorney Agent or FirmWeaver Austin Villeneuve
`Sampson LLP
`
`57
`
`ABSTRACT
`
`Methods and apparatus for genorrie analysis are provided A
`microfahricated structure including a microfluidic distribu
`tion channel
`is configured to distribute microreactor elements
`having copies of a sequencing template into a plurality of
`micrafabricated thennal cycling chambers A microreactor
`a microcarrier element carrying the
`element may include
`multiple copies of tlie sequencing template The mi era ca rrier
`element may comprise a microThere An autovalve at an exit
`port of a thermal cycling chamber an optical
`scanner or a
`timing arrangement may be used to ensure that only one
`flow into one thermal cycling chamber
`microsphere will
`wherein thermal cycling extension fragments
`are produced
`The extension products are captured purified and concen
`ide gel capture chamber A
`trated in an integrated oligonucieot
`separation apparatus is used to
`microfabricated component
`fragments The microfabri
`analyze the purified extension
`cated structure may be used in a process for performing
`sequencing and other genetic analysis of DNA or RNA
`
`Continued
`
`16 Claims 12 Drawing Sheets
`
`Thermal mline chamber
`
`Diehlwarm channel
`g16
`
`90
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`Page 1 of 29
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`BioRad Exhibit 2007
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`Page 2 of 29
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`

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`IPR201700055
`Page 3 of 29
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`

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`Structures
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`Apparatus
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`2005 510 8 pages
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`
`cited by examiner
`
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`US Patent
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`Sep 21 2010
`
`Sheet
`
`1 of 12
`
`US 7799553 B2
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`100
`
`Conventional
`
`1Arst
`
`o2
`
`00OOcOO¼QO
`
`Shear
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`Vector
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`Ligate
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`Transform
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`extract DNA
`1
`30 million timesI
`
`01
`
`1
`
`i
`
`Sanger
`extension
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`Electrophoresis
`
`FIG I
`
`11
`
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`102
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`104
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`106
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`108
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`110
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`112
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`114
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`

`

`US Patent
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`Sep 21 2010
`
`Sheet 2 of 12
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`US 7799553 B2
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`MINDS
`
`cs
`
`NTpqroN
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`41
`
`Shear
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`202
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`206
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`206
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`212
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`
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`FIG 2
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`Sep 21 2010
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`Sheet 3 of 12
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`US 7799553 B2
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`300
`
`Isolate genomic DNA
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`302
`
`Shear DNA
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`3°4
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`Enzymatic treatment
`
`306
`
`Victor Ligate
`
`308
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`Ligation mixture
`
`31°
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`100
`
`50
`
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`Exonuclease digestion
`
`312
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`Insert Recircularized
`
`FIG 3
`
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`US Patent
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`Sep 21 2010
`
`Sheet 4 of 12
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`US 7799553 B2
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`FIG 4B
`
`FIG 4A
`
`400
`
`412
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`US Patent
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`Sep 21 2010
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`Sheet 5 of 12
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`US 7799553 B2
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`405
`
`FIG 4C
`
`432
`
`436
`
`434
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`US Patent
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`Sep 21 2010
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`Sheet 6 of 12
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`US 7799553 B2
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`500
`
`522
`
`500
`
`522
`
`500
`
`522
`
`FIG 5A
`
`FIG 5B
`
`FIG 5C
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`US Patent
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`Sep 21 2010
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`Sheet 7 of 12
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`US 7799553 B2
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`608
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`FIG GA
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`608
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`602
`
`FIG GB
`
`607
`
`602
`
`606
`
`608
`
`600 FIG 6C
`
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`US Patent
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`Sep 21 2010
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`Sheet 8 of 12
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`US 7799553 B2
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`708
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`Constriction
`
`FIG 7A
`
`FIG 7B
`
`Micro sphere
`
`700
`
`702
`
`708
`
`702
`
`708
`
`702
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`US Patent
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`Sep 21 2010
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`Sheet 9 of 12
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`US 7799553 B2
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`802
`
`8B
`
`808
`
`807
`
`8B
`
`FIG 8A
`
`808
`
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`
`FIG 8B
`
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`US Patent
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`Sep 21 2010
`
`Sheet 10 412
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`US 7799553 B2
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`Thermal cycling chamber 920
`RIDS
`
`900
`
`902
`
`904
`
`918b
`
`908
`
`906
`
`Sample purihcation chamber
`
`Distribution channel
`
`904
`
`Heaters FIG 9A
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`902
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`Thermal cycling chamber
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`Distribution channel
`
`926
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`
`922
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`Atir
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`
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`918b
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`X
`
`Sample purification
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`
`924
`
`Separation channels
`
`FIG 9B
`
`908
`
`BioRad Exhibit 2007
`