JP5160433B2 - 迅速並行核酸分析 - Google Patents
迅速並行核酸分析 Download PDFInfo
- Publication number
- JP5160433B2 JP5160433B2 JP2008536859A JP2008536859A JP5160433B2 JP 5160433 B2 JP5160433 B2 JP 5160433B2 JP 2008536859 A JP2008536859 A JP 2008536859A JP 2008536859 A JP2008536859 A JP 2008536859A JP 5160433 B2 JP5160433 B2 JP 5160433B2
- Authority
- JP
- Japan
- Prior art keywords
- nucleic acid
- template
- primer
- nucleotide
- nucleotides
- Prior art date
- Legal status (The legal status is an assumption and is not a legal conclusion. Google has not performed a legal analysis and makes no representation as to the accuracy of the status listed.)
- Active
Links
- 150000007523 nucleic acids Chemical class 0.000 title claims description 140
- 108020004707 nucleic acids Proteins 0.000 title claims description 135
- 102000039446 nucleic acids Human genes 0.000 title claims description 135
- 238000004458 analytical method Methods 0.000 title description 2
- 239000002773 nucleotide Substances 0.000 claims description 222
- 125000003729 nucleotide group Chemical group 0.000 claims description 222
- 238000000034 method Methods 0.000 claims description 119
- 238000006243 chemical reaction Methods 0.000 claims description 66
- 239000011541 reaction mixture Substances 0.000 claims description 60
- 102000004190 Enzymes Human genes 0.000 claims description 48
- 108090000790 Enzymes Proteins 0.000 claims description 48
- 238000006116 polymerization reaction Methods 0.000 claims description 48
- 238000001514 detection method Methods 0.000 claims description 42
- 238000012163 sequencing technique Methods 0.000 claims description 39
- 229910019142 PO4 Inorganic materials 0.000 claims description 38
- 239000000975 dye Substances 0.000 claims description 37
- 239000010452 phosphate Substances 0.000 claims description 37
- 150000001768 cations Chemical class 0.000 claims description 36
- 239000011324 bead Substances 0.000 claims description 35
- 230000000379 polymerizing effect Effects 0.000 claims description 32
- 108010014303 DNA-directed DNA polymerase Proteins 0.000 claims description 30
- 102000016928 DNA-directed DNA polymerase Human genes 0.000 claims description 30
- 230000000295 complement effect Effects 0.000 claims description 29
- 125000002467 phosphate group Chemical group [H]OP(=O)(O[H])O[*] 0.000 claims description 27
- 229920000388 Polyphosphate Polymers 0.000 claims description 17
- 239000001205 polyphosphate Substances 0.000 claims description 17
- 235000011176 polyphosphates Nutrition 0.000 claims description 17
- XPPKVPWEQAFLFU-UHFFFAOYSA-J diphosphate(4-) Chemical group [O-]P([O-])(=O)OP([O-])([O-])=O XPPKVPWEQAFLFU-UHFFFAOYSA-J 0.000 claims description 13
- 235000011180 diphosphates Nutrition 0.000 claims description 13
- 108060001084 Luciferase Proteins 0.000 claims description 12
- 239000005089 Luciferase Substances 0.000 claims description 12
- 239000011777 magnesium Substances 0.000 claims description 11
- -1 Klenow exo Proteins 0.000 claims description 10
- 238000005755 formation reaction Methods 0.000 claims description 10
- 230000008569 process Effects 0.000 claims description 10
- 230000009918 complex formation Effects 0.000 claims description 8
- ZYGHJZDHTFUPRJ-UHFFFAOYSA-N coumarin Chemical compound C1=CC=C2OC(=O)C=CC2=C1 ZYGHJZDHTFUPRJ-UHFFFAOYSA-N 0.000 claims description 8
- 108010092799 RNA-directed DNA polymerase Proteins 0.000 claims description 6
- 239000006227 byproduct Substances 0.000 claims description 6
- 239000002738 chelating agent Substances 0.000 claims description 6
- 239000007850 fluorescent dye Substances 0.000 claims description 6
- 102100034343 Integrase Human genes 0.000 claims description 5
- 102000004523 Sulfate Adenylyltransferase Human genes 0.000 claims description 4
- 108010022348 Sulfate adenylyltransferase Proteins 0.000 claims description 4
- 239000000987 azo dye Substances 0.000 claims description 4
- 229960000956 coumarin Drugs 0.000 claims description 4
- 235000001671 coumarin Nutrition 0.000 claims description 4
- 150000004032 porphyrins Chemical class 0.000 claims description 4
- ANRHNWWPFJCPAZ-UHFFFAOYSA-M thionine Chemical compound [Cl-].C1=CC(N)=CC2=[S+]C3=CC(N)=CC=C3N=C21 ANRHNWWPFJCPAZ-UHFFFAOYSA-M 0.000 claims description 4
- 239000001018 xanthene dye Substances 0.000 claims description 4
- 102000004594 DNA Polymerase I Human genes 0.000 claims description 3
- 108010017826 DNA Polymerase I Proteins 0.000 claims description 3
- FYYHWMGAXLPEAU-UHFFFAOYSA-N Magnesium Chemical compound [Mg] FYYHWMGAXLPEAU-UHFFFAOYSA-N 0.000 claims description 3
- 229910052749 magnesium Inorganic materials 0.000 claims description 3
- 239000011572 manganese Substances 0.000 claims description 3
- DZVCFNFOPIZQKX-LTHRDKTGSA-M merocyanine Chemical compound [Na+].O=C1N(CCCC)C(=O)N(CCCC)C(=O)C1=C\C=C\C=C/1N(CCCS([O-])(=O)=O)C2=CC=CC=C2O\1 DZVCFNFOPIZQKX-LTHRDKTGSA-M 0.000 claims description 3
- PWHULOQIROXLJO-UHFFFAOYSA-N Manganese Chemical compound [Mn] PWHULOQIROXLJO-UHFFFAOYSA-N 0.000 claims description 2
- 102000004160 Phosphoric Monoester Hydrolases Human genes 0.000 claims description 2
- 108090000608 Phosphoric Monoester Hydrolases Proteins 0.000 claims description 2
- 230000003100 immobilizing effect Effects 0.000 claims description 2
- 229910052748 manganese Inorganic materials 0.000 claims description 2
- 108010058966 bacteriophage T7 induced DNA polymerase Proteins 0.000 claims 4
- 239000000835 fiber Substances 0.000 claims 2
- 101100409165 Escherichia coli (strain K12) prc gene Proteins 0.000 claims 1
- 108060002716 Exonuclease Proteins 0.000 claims 1
- 108010078851 HIV Reverse Transcriptase Proteins 0.000 claims 1
- 101000902539 Homo sapiens DNA polymerase beta Proteins 0.000 claims 1
- WAEMQWOKJMHJLA-UHFFFAOYSA-N Manganese(2+) Chemical compound [Mn+2] WAEMQWOKJMHJLA-UHFFFAOYSA-N 0.000 claims 1
- 241000713869 Moloney murine leukemia virus Species 0.000 claims 1
- 108010006785 Taq Polymerase Proteins 0.000 claims 1
- 241000074588 Thermogutta hypogea Species 0.000 claims 1
- 230000002950 deficient Effects 0.000 claims 1
- 102000013165 exonuclease Human genes 0.000 claims 1
- 239000011521 glass Substances 0.000 claims 1
- 102000047799 human POLB Human genes 0.000 claims 1
- 235000021317 phosphate Nutrition 0.000 description 35
- 108020004414 DNA Proteins 0.000 description 23
- 238000012175 pyrosequencing Methods 0.000 description 19
- 239000002777 nucleoside Substances 0.000 description 13
- 239000000047 product Substances 0.000 description 11
- PEDCQBHIVMGVHV-UHFFFAOYSA-N Glycerine Chemical compound OCC(O)CO PEDCQBHIVMGVHV-UHFFFAOYSA-N 0.000 description 9
- 108091028043 Nucleic acid sequence Proteins 0.000 description 9
- 230000015572 biosynthetic process Effects 0.000 description 9
- 239000007795 chemical reaction product Substances 0.000 description 9
- KCXVZYZYPLLWCC-UHFFFAOYSA-N EDTA Chemical compound OC(=O)CN(CC(O)=O)CCN(CC(O)=O)CC(O)=O KCXVZYZYPLLWCC-UHFFFAOYSA-N 0.000 description 8
- NBIIXXVUZAFLBC-UHFFFAOYSA-K phosphate Chemical compound [O-]P([O-])([O-])=O NBIIXXVUZAFLBC-UHFFFAOYSA-K 0.000 description 8
- FWMNVWWHGCHHJJ-SKKKGAJSSA-N 4-amino-1-[(2r)-6-amino-2-[[(2r)-2-[[(2r)-2-[[(2r)-2-amino-3-phenylpropanoyl]amino]-3-phenylpropanoyl]amino]-4-methylpentanoyl]amino]hexanoyl]piperidine-4-carboxylic acid Chemical compound C([C@H](C(=O)N[C@H](CC(C)C)C(=O)N[C@H](CCCCN)C(=O)N1CCC(N)(CC1)C(O)=O)NC(=O)[C@H](N)CC=1C=CC=CC=1)C1=CC=CC=C1 FWMNVWWHGCHHJJ-SKKKGAJSSA-N 0.000 description 7
- 238000013500 data storage Methods 0.000 description 7
- 238000005516 engineering process Methods 0.000 description 7
- 150000003833 nucleoside derivatives Chemical class 0.000 description 7
- OAKPWEUQDVLTCN-NKWVEPMBSA-N 2',3'-Dideoxyadenosine-5-triphosphate Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@H]1CC[C@@H](CO[P@@](O)(=O)O[P@](O)(=O)OP(O)(O)=O)O1 OAKPWEUQDVLTCN-NKWVEPMBSA-N 0.000 description 6
- BTBUEUYNUDRHOZ-UHFFFAOYSA-N Borate Chemical compound [O-]B([O-])[O-] BTBUEUYNUDRHOZ-UHFFFAOYSA-N 0.000 description 6
- 239000007983 Tris buffer Substances 0.000 description 6
- 230000003321 amplification Effects 0.000 description 6
- 238000010586 diagram Methods 0.000 description 6
- 229910021645 metal ion Inorganic materials 0.000 description 6
- 238000003199 nucleic acid amplification method Methods 0.000 description 6
- 108091033319 polynucleotide Proteins 0.000 description 6
- 102000040430 polynucleotide Human genes 0.000 description 6
- 239000002157 polynucleotide Substances 0.000 description 6
- LENZDBCJOHFCAS-UHFFFAOYSA-N tris Chemical compound OCC(N)(CO)CO LENZDBCJOHFCAS-UHFFFAOYSA-N 0.000 description 6
- 238000005406 washing Methods 0.000 description 6
- 102000004163 DNA-directed RNA polymerases Human genes 0.000 description 5
- 108090000626 DNA-directed RNA polymerases Proteins 0.000 description 5
- SUYVUBYJARFZHO-RRKCRQDMSA-N dATP Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@H]1C[C@H](O)[C@@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)O1 SUYVUBYJARFZHO-RRKCRQDMSA-N 0.000 description 5
- SUYVUBYJARFZHO-UHFFFAOYSA-N dATP Natural products C1=NC=2C(N)=NC=NC=2N1C1CC(O)C(COP(O)(=O)OP(O)(=O)OP(O)(O)=O)O1 SUYVUBYJARFZHO-UHFFFAOYSA-N 0.000 description 5
- 239000000839 emulsion Substances 0.000 description 5
- 239000011325 microbead Substances 0.000 description 5
- 230000000284 resting effect Effects 0.000 description 5
- 239000000243 solution Substances 0.000 description 5
- 108091034117 Oligonucleotide Proteins 0.000 description 4
- 230000003197 catalytic effect Effects 0.000 description 4
- 238000002372 labelling Methods 0.000 description 4
- 238000004020 luminiscence type Methods 0.000 description 4
- 239000000376 reactant Substances 0.000 description 4
- 239000001226 triphosphate Substances 0.000 description 4
- 235000011178 triphosphate Nutrition 0.000 description 4
- FGUUSXIOTUKUDN-IBGZPJMESA-N C1(=CC=CC=C1)N1C2=C(NC([C@H](C1)NC=1OC(=NN=1)C1=CC=CC=C1)=O)C=CC=C2 Chemical compound C1(=CC=CC=C1)N1C2=C(NC([C@H](C1)NC=1OC(=NN=1)C1=CC=CC=C1)=O)C=CC=C2 FGUUSXIOTUKUDN-IBGZPJMESA-N 0.000 description 3
- 238000003556 assay Methods 0.000 description 3
- 230000008901 benefit Effects 0.000 description 3
- 239000000872 buffer Substances 0.000 description 3
- 230000008859 change Effects 0.000 description 3
- 230000000694 effects Effects 0.000 description 3
- 238000006911 enzymatic reaction Methods 0.000 description 3
- 125000002887 hydroxy group Chemical group [H]O* 0.000 description 3
- 238000011065 in-situ storage Methods 0.000 description 3
- 238000009790 rate-determining step (RDS) Methods 0.000 description 3
- 238000000926 separation method Methods 0.000 description 3
- YBJHBAHKTGYVGT-ZKWXMUAHSA-N (+)-Biotin Chemical compound N1C(=O)N[C@@H]2[C@H](CCCCC(=O)O)SC[C@@H]21 YBJHBAHKTGYVGT-ZKWXMUAHSA-N 0.000 description 2
- GNFTZDOKVXKIBK-UHFFFAOYSA-N 3-(2-methoxyethoxy)benzohydrazide Chemical compound COCCOC1=CC=CC(C(=O)NN)=C1 GNFTZDOKVXKIBK-UHFFFAOYSA-N 0.000 description 2
- PEHVGBZKEYRQSX-UHFFFAOYSA-N 7-deaza-adenine Chemical compound NC1=NC=NC2=C1C=CN2 PEHVGBZKEYRQSX-UHFFFAOYSA-N 0.000 description 2
- KDCGOANMDULRCW-UHFFFAOYSA-N 7H-purine Chemical compound N1=CNC2=NC=NC2=C1 KDCGOANMDULRCW-UHFFFAOYSA-N 0.000 description 2
- IGXWBGJHJZYPQS-SSDOTTSWSA-N D-Luciferin Chemical compound OC(=O)[C@H]1CSC(C=2SC3=CC=C(O)C=C3N=2)=N1 IGXWBGJHJZYPQS-SSDOTTSWSA-N 0.000 description 2
- CYCGRDQQIOGCKX-UHFFFAOYSA-N Dehydro-luciferin Natural products OC(=O)C1=CSC(C=2SC3=CC(O)=CC=C3N=2)=N1 CYCGRDQQIOGCKX-UHFFFAOYSA-N 0.000 description 2
- BJGNCJDXODQBOB-UHFFFAOYSA-N Fivefly Luciferin Natural products OC(=O)C1CSC(C=2SC3=CC(O)=CC=C3N=2)=N1 BJGNCJDXODQBOB-UHFFFAOYSA-N 0.000 description 2
- DDWFXDSYGUXRAY-UHFFFAOYSA-N Luciferin Natural products CCc1c(C)c(CC2NC(=O)C(=C2C=C)C)[nH]c1Cc3[nH]c4C(=C5/NC(CC(=O)O)C(C)C5CC(=O)O)CC(=O)c4c3C DDWFXDSYGUXRAY-UHFFFAOYSA-N 0.000 description 2
- 206010029719 Nonspecific reaction Diseases 0.000 description 2
- IQFYYKKMVGJFEH-XLPZGREQSA-N Thymidine Chemical group O=C1NC(=O)C(C)=CN1[C@@H]1O[C@H](CO)[C@@H](O)C1 IQFYYKKMVGJFEH-XLPZGREQSA-N 0.000 description 2
- ISAKRJDGNUQOIC-UHFFFAOYSA-N Uracil Chemical compound O=C1C=CNC(=O)N1 ISAKRJDGNUQOIC-UHFFFAOYSA-N 0.000 description 2
- OIRDTQYFTABQOQ-KQYNXXCUSA-N adenosine Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@H](CO)[C@@H](O)[C@H]1O OIRDTQYFTABQOQ-KQYNXXCUSA-N 0.000 description 2
- IRLPACMLTUPBCL-FCIPNVEPSA-N adenosine-5'-phosphosulfate Chemical compound C1=NC=2C(N)=NC=NC=2N1[C@@H]1O[C@@H](CO[P@](O)(=O)OS(O)(=O)=O)[C@H](O)[C@H]1O IRLPACMLTUPBCL-FCIPNVEPSA-N 0.000 description 2
- OPTASPLRGRRNAP-UHFFFAOYSA-N cytosine Chemical compound NC=1C=CNC(=O)N=1 OPTASPLRGRRNAP-UHFFFAOYSA-N 0.000 description 2
- 238000001962 electrophoresis Methods 0.000 description 2
- LIYGYAHYXQDGEP-UHFFFAOYSA-N firefly oxyluciferin Natural products Oc1csc(n1)-c1nc2ccc(O)cc2s1 LIYGYAHYXQDGEP-UHFFFAOYSA-N 0.000 description 2
- UYTPUPDQBNUYGX-UHFFFAOYSA-N guanine Chemical compound O=C1NC(N)=NC2=C1N=CN2 UYTPUPDQBNUYGX-UHFFFAOYSA-N 0.000 description 2
- FDGQSTZJBFJUBT-UHFFFAOYSA-N hypoxanthine Chemical compound O=C1NC=NC2=C1NC=N2 FDGQSTZJBFJUBT-UHFFFAOYSA-N 0.000 description 2
- 235000019689 luncheon sausage Nutrition 0.000 description 2
- 238000005259 measurement Methods 0.000 description 2
- 238000001668 nucleic acid synthesis Methods 0.000 description 2
- JJVOROULKOMTKG-UHFFFAOYSA-N oxidized Photinus luciferin Chemical compound S1C2=CC(O)=CC=C2N=C1C1=NC(=O)CS1 JJVOROULKOMTKG-UHFFFAOYSA-N 0.000 description 2
- 230000036961 partial effect Effects 0.000 description 2
- 238000011160 research Methods 0.000 description 2
- 241000894007 species Species 0.000 description 2
- 239000000126 substance Substances 0.000 description 2
- RWQNBRDOKXIBIV-UHFFFAOYSA-N thymine Chemical compound CC1=CNC(=O)NC1=O RWQNBRDOKXIBIV-UHFFFAOYSA-N 0.000 description 2
- 238000004448 titration Methods 0.000 description 2
- 125000002264 triphosphate group Chemical class [H]OP(=O)(O[H])OP(=O)(O[H])OP(=O)(O[H])O* 0.000 description 2
- YKBGVTZYEHREMT-KVQBGUIXSA-N 2'-deoxyguanosine Chemical compound C1=NC=2C(=O)NC(N)=NC=2N1[C@H]1C[C@H](O)[C@@H](CO)O1 YKBGVTZYEHREMT-KVQBGUIXSA-N 0.000 description 1
- CKTSBUTUHBMZGZ-SHYZEUOFSA-N 2'‐deoxycytidine Chemical compound O=C1N=C(N)C=CN1[C@@H]1O[C@H](CO)[C@@H](O)C1 CKTSBUTUHBMZGZ-SHYZEUOFSA-N 0.000 description 1
- LRSASMSXMSNRBT-UHFFFAOYSA-N 5-methylcytosine Chemical compound CC1=CNC(=O)N=C1N LRSASMSXMSNRBT-UHFFFAOYSA-N 0.000 description 1
- LOSIULRWFAEMFL-UHFFFAOYSA-N 7-deazaguanine Chemical compound O=C1NC(N)=NC2=C1CC=N2 LOSIULRWFAEMFL-UHFFFAOYSA-N 0.000 description 1
- MSSXOMSJDRHRMC-UHFFFAOYSA-N 9H-purine-2,6-diamine Chemical compound NC1=NC(N)=C2NC=NC2=N1 MSSXOMSJDRHRMC-UHFFFAOYSA-N 0.000 description 1
- 229930024421 Adenine Natural products 0.000 description 1
- GFFGJBXGBJISGV-UHFFFAOYSA-N Adenine Chemical compound NC1=NC=NC2=C1N=CN2 GFFGJBXGBJISGV-UHFFFAOYSA-N 0.000 description 1
- 108091093088 Amplicon Proteins 0.000 description 1
- DWRXFEITVBNRMK-UHFFFAOYSA-N Beta-D-1-Arabinofuranosylthymine Natural products O=C1NC(=O)C(C)=CN1C1C(O)C(O)C(CO)O1 DWRXFEITVBNRMK-UHFFFAOYSA-N 0.000 description 1
- 108020004635 Complementary DNA Proteins 0.000 description 1
- 102000053602 DNA Human genes 0.000 description 1
- 102000010567 DNA Polymerase II Human genes 0.000 description 1
- 108010063113 DNA Polymerase II Proteins 0.000 description 1
- 102000007528 DNA Polymerase III Human genes 0.000 description 1
- 108010071146 DNA Polymerase III Proteins 0.000 description 1
- 102000001996 DNA Polymerase beta Human genes 0.000 description 1
- 108010001132 DNA Polymerase beta Proteins 0.000 description 1
- 102000016903 DNA Polymerase gamma Human genes 0.000 description 1
- 108010014080 DNA Polymerase gamma Proteins 0.000 description 1
- 108010092681 DNA Primase Proteins 0.000 description 1
- 102000016559 DNA Primase Human genes 0.000 description 1
- 108010008286 DNA nucleotidylexotransferase Proteins 0.000 description 1
- 102100033215 DNA nucleotidylexotransferase Human genes 0.000 description 1
- 238000001712 DNA sequencing Methods 0.000 description 1
- 230000006820 DNA synthesis Effects 0.000 description 1
- CKTSBUTUHBMZGZ-UHFFFAOYSA-N Deoxycytidine Natural products O=C1N=C(N)C=CN1C1OC(CO)C(O)C1 CKTSBUTUHBMZGZ-UHFFFAOYSA-N 0.000 description 1
- UGQMRVRMYYASKQ-UHFFFAOYSA-N Hypoxanthine nucleoside Natural products OC1C(O)C(CO)OC1N1C(NC=NC2=O)=C2N=C1 UGQMRVRMYYASKQ-UHFFFAOYSA-N 0.000 description 1
- JLVVSXFLKOJNIY-UHFFFAOYSA-N Magnesium ion Chemical compound [Mg+2] JLVVSXFLKOJNIY-UHFFFAOYSA-N 0.000 description 1
- CZPWVGJYEJSRLH-UHFFFAOYSA-N Pyrimidine Chemical compound C1=CN=CN=C1 CZPWVGJYEJSRLH-UHFFFAOYSA-N 0.000 description 1
- 108020004511 Recombinant DNA Proteins 0.000 description 1
- 229920002684 Sepharose Polymers 0.000 description 1
- 108020004682 Single-Stranded DNA Proteins 0.000 description 1
- 108010090804 Streptavidin Proteins 0.000 description 1
- 108010017842 Telomerase Proteins 0.000 description 1
- YTAHJIFKAKIKAV-XNMGPUDCSA-N [(1R)-3-morpholin-4-yl-1-phenylpropyl] N-[(3S)-2-oxo-5-phenyl-1,3-dihydro-1,4-benzodiazepin-3-yl]carbamate Chemical compound O=C1[C@H](N=C(C2=C(N1)C=CC=C2)C1=CC=CC=C1)NC(O[C@H](CCN1CCOCC1)C1=CC=CC=C1)=O YTAHJIFKAKIKAV-XNMGPUDCSA-N 0.000 description 1
- JLCPHMBAVCMARE-UHFFFAOYSA-N [3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-[[3-[[3-[[3-[[3-[[3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-[[5-(2-amino-6-oxo-1H-purin-9-yl)-3-hydroxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxyoxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(5-methyl-2,4-dioxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(6-aminopurin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl]oxy-5-(4-amino-2-oxopyrimidin-1-yl)oxolan-2-yl]methyl [5-(6-aminopurin-9-yl)-2-(hydroxymethyl)oxolan-3-yl] hydrogen phosphate Polymers Cc1cn(C2CC(OP(O)(=O)OCC3OC(CC3OP(O)(=O)OCC3OC(CC3O)n3cnc4c3nc(N)[nH]c4=O)n3cnc4c3nc(N)[nH]c4=O)C(COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3COP(O)(=O)OC3CC(OC3CO)n3cnc4c(N)ncnc34)n3ccc(N)nc3=O)n3cnc4c(N)ncnc34)n3ccc(N)nc3=O)n3ccc(N)nc3=O)n3ccc(N)nc3=O)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cc(C)c(=O)[nH]c3=O)n3cc(C)c(=O)[nH]c3=O)n3ccc(N)nc3=O)n3cc(C)c(=O)[nH]c3=O)n3cnc4c3nc(N)[nH]c4=O)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)n3cnc4c(N)ncnc34)O2)c(=O)[nH]c1=O JLCPHMBAVCMARE-UHFFFAOYSA-N 0.000 description 1
- HDRRAMINWIWTNU-NTSWFWBYSA-N [[(2s,5r)-5-(2-amino-6-oxo-3h-purin-9-yl)oxolan-2-yl]methoxy-hydroxyphosphoryl] phosphono hydrogen phosphate Chemical compound C1=2NC(N)=NC(=O)C=2N=CN1[C@H]1CC[C@@H](COP(O)(=O)OP(O)(=O)OP(O)(O)=O)O1 HDRRAMINWIWTNU-NTSWFWBYSA-N 0.000 description 1
- 125000002015 acyclic group Chemical group 0.000 description 1
- 229960000643 adenine Drugs 0.000 description 1
- IQFYYKKMVGJFEH-UHFFFAOYSA-N beta-L-thymidine Natural products O=C1NC(=O)C(C)=CN1C1OC(CO)C(O)C1 IQFYYKKMVGJFEH-UHFFFAOYSA-N 0.000 description 1
- 229960002685 biotin Drugs 0.000 description 1
- 235000020958 biotin Nutrition 0.000 description 1
- 239000011616 biotin Substances 0.000 description 1
- 125000002837 carbocyclic group Chemical group 0.000 description 1
- 238000006555 catalytic reaction Methods 0.000 description 1
- 238000001311 chemical methods and process Methods 0.000 description 1
- 238000003776 cleavage reaction Methods 0.000 description 1
- 239000011248 coating agent Substances 0.000 description 1
- 238000000576 coating method Methods 0.000 description 1
- 238000004040 coloring Methods 0.000 description 1
- 150000001875 compounds Chemical class 0.000 description 1
- 238000012258 culturing Methods 0.000 description 1
- 229940104302 cytosine Drugs 0.000 description 1
- 230000007423 decrease Effects 0.000 description 1
- 238000004925 denaturation Methods 0.000 description 1
- 230000036425 denaturation Effects 0.000 description 1
- 239000005549 deoxyribonucleoside Substances 0.000 description 1
- 230000001419 dependent effect Effects 0.000 description 1
- 230000032050 esterification Effects 0.000 description 1
- 238000005886 esterification reaction Methods 0.000 description 1
- 238000002474 experimental method Methods 0.000 description 1
- 230000007274 generation of a signal involved in cell-cell signaling Effects 0.000 description 1
- 230000001771 impaired effect Effects 0.000 description 1
- 238000010348 incorporation Methods 0.000 description 1
- 238000011534 incubation Methods 0.000 description 1
- 230000002452 interceptive effect Effects 0.000 description 1
- 229910001425 magnesium ion Inorganic materials 0.000 description 1
- 239000000463 material Substances 0.000 description 1
- 230000004048 modification Effects 0.000 description 1
- 238000012986 modification Methods 0.000 description 1
- 238000012544 monitoring process Methods 0.000 description 1
- 238000010534 nucleophilic substitution reaction Methods 0.000 description 1
- 125000003835 nucleoside group Chemical group 0.000 description 1
- 239000013307 optical fiber Substances 0.000 description 1
- 125000001805 pentosyl group Chemical group 0.000 description 1
- 150000003013 phosphoric acid derivatives Chemical group 0.000 description 1
- 229920002401 polyacrylamide Polymers 0.000 description 1
- 238000003752 polymerase chain reaction Methods 0.000 description 1
- 230000037452 priming Effects 0.000 description 1
- 230000008707 rearrangement Effects 0.000 description 1
- 230000010076 replication Effects 0.000 description 1
- 239000002342 ribonucleoside Substances 0.000 description 1
- 238000005096 rolling process Methods 0.000 description 1
- 230000007017 scission Effects 0.000 description 1
- 238000011896 sensitive detection Methods 0.000 description 1
- 230000035945 sensitivity Effects 0.000 description 1
- 239000007787 solid Substances 0.000 description 1
- 238000006467 substitution reaction Methods 0.000 description 1
- 235000021092 sugar substitutes Nutrition 0.000 description 1
- 239000000725 suspension Substances 0.000 description 1
- 239000003765 sweetening agent Substances 0.000 description 1
- 238000003786 synthesis reaction Methods 0.000 description 1
- 229940104230 thymidine Drugs 0.000 description 1
- 229940113082 thymine Drugs 0.000 description 1
- 229940035893 uracil Drugs 0.000 description 1
Classifications
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6869—Methods for sequencing
-
- C—CHEMISTRY; METALLURGY
- C12—BIOCHEMISTRY; BEER; SPIRITS; WINE; VINEGAR; MICROBIOLOGY; ENZYMOLOGY; MUTATION OR GENETIC ENGINEERING
- C12Q—MEASURING OR TESTING PROCESSES INVOLVING ENZYMES, NUCLEIC ACIDS OR MICROORGANISMS; COMPOSITIONS OR TEST PAPERS THEREFOR; PROCESSES OF PREPARING SUCH COMPOSITIONS; CONDITION-RESPONSIVE CONTROL IN MICROBIOLOGICAL OR ENZYMOLOGICAL PROCESSES
- C12Q1/00—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions
- C12Q1/68—Measuring or testing processes involving enzymes, nucleic acids or microorganisms; Compositions therefor; Processes of preparing such compositions involving nucleic acids
- C12Q1/6869—Methods for sequencing
- C12Q1/6874—Methods for sequencing involving nucleic acid arrays, e.g. sequencing by hybridisation
Description
「ヌクレオシド」という用語は、本発明では、プリン、デアザプリン、ピリミジン、又は修飾塩基が、糖又は糖代替物、例えば炭素環式又は非環式部分に、1’位又は同等の位置で結合したものを含む化合物のことを意味するものであり、2’−デオキシ及び2’−ヒドロキシル、及び2’,3’−ジデオキシ体、ならびに他の置換体も含む。
「オリゴヌクレオチド」という用語は、ヌクレオチド又はその誘導体の線状オリゴマー、例えばデオキシリボヌクレオシド、リボヌクレオシドなどを包含する。本明細書では、特記しない限り、オリゴヌクレオチドをアルファベットの配列で表す場合には、ヌクレオチドは、左から右に5’→3’の順序で表示し、Aは、デオキシアデノシンを、Cはデオキシシチジンを、Gはデオキシグアノシンを、そしてTはチミジンを示す。
ヌクレオチドの取り込みというイベントを検出するもう一つの方法は、ヌクレオチドが成長中の鎖に取り込まれた後に、反応副生物を検出する方法である。ヌクレオチドは、末端の燐酸上に、酵素で活性化が可能な標識を含むことができる。重合後、この酵素で活性化が可能な標識酵素は、無機ポリリン酸副生物上に位置することになる。ホスファターゼによってポリリン酸副生物を開裂させると、検出可能な標識を生成することができる。各ヌクレオチドごとに別々の標識を用いる場合には、4種すべてのヌクレオチドを、同じ反応で区別することができる。別々の標識を使用しない場合には、各ヌクレオチドを別々に導入して、異なるヌクレオチドを識別する。あるいは、重合反応の間に非標識ヌクレオチドから放出されるPPiを、各種の検定法で検出することもできるが、この場合、各塩基を区別するために、この工程を繰り返すことが必要になる。こうした検定法の1種は、一連の反応カスケードを通じて光子の放出を検出することによって実施される。この方法では、まず、ATPスルフリラーゼを使用してPPiとアデノシン−5’−ホスホ硫酸からATPを生成し、次に、ルシフェラーゼによってATPとルシフェリンをオキシルシフェリンに変換して、光子を発生させる(Nyren et al., 151 Analytical Biochemistry 504 (1985))。
図5は、蛍光標識ヌクレオチドを使用することによって、この種の安定した閉鎖複合体が実際に形成されていることを示す。この図には、本発明に記載した方法によって、複合体を検出できることが明瞭に示されている。ポリメラーゼ反応(20ul)を、50pmoleの図中に示すプライマー結合鋳型、±20pmoleの正荷電標識ddGTP及び/又はddATP、±3pmoleのFY7DNAポリメラーゼを含む(25mMのTris:Borate、pH=7.5、0.1mM EDTA、10%グリセロール)中で実施した。反応生成物を、50mMのTris:Borate(pH=7.5)へのPAGEの7%溶液中で分離した。複合体の形成は、ポリメラーゼ、プライマー鋳型、正しいヌクレオチドが存在する場合のみ観察された。
Claims (27)
- 複数の核酸鋳型の配列を並行して決定する方法であって、
a)複数のプライマー結合核酸鋳型を支持構造体に固定化する工程であって、各プライマー結合核酸鋳型が鋳型とプライマーとを含んでいて、上記支持構造体の識別可能な離散した位置に局在化している工程、
b)上記支持構造体上で、反応混合物を形成することによって、複数の複合体形成反応を開始する工程であって、上記反応混合物が、上記複数のプライマー結合核酸鋳型と、核酸重合酵素と、1種以上のヌクレオチドとを含んでいて複数の閉鎖複合体を形成し、閉鎖複合体の各々が、上記複数のプライマー結合核酸鋳型のいずれかと、上記核酸重合酵素と、重合部位における鋳型と相補的な塩基を含む1種以上のヌクレオチドとを含んでいる、工程、
c)上記反応混合物中の非結合ヌクレオチド及び他の非結合成分を除去する工程、
d)マグネシウム及びマンガンから選択される二価陽イオンを加えて重合反応を完了させる工程、
e)識別可能な離散した位置の各々において各閉鎖複合体について、導入核酸を検出する工程、
f)二価陽イオンを除去する工程、及び
g)工程b)〜f)を繰り返して、複数のプライマー結合核酸鋳型の各追加ヌクレオチドの配列を決定する工程
を含んでなる方法。 - 上記支持構造体がビーズを含んでおり、該ビーズが単一のプライマーと鋳型の組合せを担持しており、別のプライマーと鋳型の組合せを担持するビーズとは識別可能に分離されている、請求項1記載の方法。
- 上記鋳型がポリメラーゼコロニー法で上記ビーズに固定化される、請求項2記載の方法。
- 上記ビーズがマルチウェルプレートの個々のウェルに支持され、ビーズが個々のウェルに分離されている、請求項2記載の方法。
- 識別可能な離散した位置が96ウェルプレートの個々のウェルであり、96以下の配列決定反応が同時に並行して実施される、請求項1記載の方法。
- 識別可能な離散した位置が384ウェルプレートの個々のウェルであり、384以下の配列決定反応が同時に並行して実施される、請求項1記載の方法。
- 識別可能な離散した位置が1536ウェルプレートの個々のウェルであり、1536以下の配列決定反応が同時に並行して実施される、請求項1記載の方法。
- 各ビーズが、光ファイバースライドのウェル内に導入することによって分離される、請求項2記載の方法。
- 前記光ファイバースライドの各ウェルが1個のビーズを保持する、請求項8記載の方法。
- 前記支持構造体がスライドガラスの第一表面である、請求項1記載の方法。
- 前記工程(b)の反応混合物がさらにキレート剤を含む、請求項1記載の方法。
- 前記核酸重合酵素がDNAポリメラーゼ又は逆転写酵素である、請求項1記載の方法。
- 前記核酸重合酵素がDNAポリメラーゼI、T4DNAポリメラーゼ、Amplitaq FS、T7DNAポリメラーゼ、Phi29DNAポリメラーゼ、クレノーエキソ、シークエナーゼ、TaqDNAポリメラーゼ、サーモシークエナーゼI、サーモシークエナーゼII、FY7 DNAポリメラーゼ、サーモシークエナーゼE681M、T.hypogea(Thy B)DNAポリメラーゼ、T.neapolitana(Tne)DNAポリメラーゼ、T.subterranea(Tsu)DNAポリメラーゼ、T.barossii(Tba)DNAポリメラーゼ、T.litoralis(Vent)DNAポリメラーゼ、T.kodakaraensis DNAポリメラーゼ、P.furiosisDNAポリメラーゼ、P.GB−D(Deep Vent)DNAポリメラーゼ、ヒトPol beta、Tsp JS1 DNAポリメラーゼ、AMV逆転写酵素、MMLV逆転写酵素、HIV逆転写酵素、これらのポリメラーゼのエキソヌクレアーゼ欠乏性変異体から選択される、請求項1記載の方法。
- 前記1種以上のヌクレオチドの末端リン酸が標識されている、請求項1記載の方法。
- 検出工程(e)が前記添加工程(d)の前に実施され、各々の異なるヌクレオチドが別々の標識を担持していて、該検出工程が、上記末端リン酸標識ヌクレオチドの別々の標識を各位置で検出することによって実施される、請求項14記載の方法。
- 1種以上のヌクレオチドの各々が4種類のヌクレオチドであり、各々別々の標識を担持していて、4種類の天然塩基のいずれかと相補的である、請求項14記載の方法。
- 前記末端リン酸標識ヌクレオチドの標識が、蛍光色素、着色色素、又は化学発光原子団である、請求項14記載の方法。
- 前記蛍光色素が、キサンテン色素、シアニン色素、メロシアニン色素、アゾ色素、ポルフィリン色素、クマリン色素、Bodipy色素及びそれらの誘導体からなる群から選択される、請求項17記載の方法。
- 前記着色色素が、アゾ色素、メロシアニン、シアニン色素、キサンテン色素、ポルフィリン色素、クマリン色素、Bodipy色素及びそれらの誘導体からなる群から選択される、請求項17記載の方法。
- 前記標識が、重合反応の無機ポリリン酸塩副生物上に存在する、酵素活性化可能な標識であり、上記検出工程が、ポリリン酸塩副生物をホスファターゼで開裂して検出可能な標識を生成する工程をさらに含む、請求項14記載の方法。
- 前記検出工程(e)が、上記ヌクレオチドから放出された無機ポリリン酸塩を測定して、ヌクレオチドの塩基の種類を推定することによって実施される、請求項1記載の方法。
- 前記検出工程が、光子の測定によって実施され、ここで、上記ポリリン酸塩がピロリン酸であってまずATPスルフリラーゼによってATPに転化され、その後、ルシフェラーゼ反応によって光子が生成される、請求項21記載の方法。
- 前記二価陽イオンがマグネシウム(Mg2+)である、請求項1記載の方法。
- 前記二価陽イオンがマンガン(Mn2+)である、請求項1記載の方法。
- 工程(a)において、プライマー結合核酸鋳型を形成してから支持構造体に固定化する、請求項1記載の方法。
- 工程(a)において、まず核酸鋳型を支持構造体に固定化してから、上記プライマーとプライマー結合核酸鋳型を形成する、請求項1記載の方法。
- 工程(a)において、まずプライマーを支持構造体に固定化してから、上記核酸鋳型とプライマー結合核酸鋳型を形成する、請求項1記載の方法。
Applications Claiming Priority (3)
Application Number | Priority Date | Filing Date | Title |
---|---|---|---|
US11/255,683 | 2005-10-21 | ||
US11/255,683 US7264934B2 (en) | 2004-06-10 | 2005-10-21 | Rapid parallel nucleic acid analysis |
PCT/US2006/041232 WO2007048033A1 (en) | 2005-10-21 | 2006-10-20 | Rapid parallel nucleic acid analysis |
Publications (2)
Publication Number | Publication Date |
---|---|
JP2009512452A JP2009512452A (ja) | 2009-03-26 |
JP5160433B2 true JP5160433B2 (ja) | 2013-03-13 |
Family
ID=37962856
Family Applications (1)
Application Number | Title | Priority Date | Filing Date |
---|---|---|---|
JP2008536859A Active JP5160433B2 (ja) | 2005-10-21 | 2006-10-20 | 迅速並行核酸分析 |
Country Status (4)
Country | Link |
---|---|
US (1) | US7264934B2 (ja) |
EP (1) | EP1954827B1 (ja) |
JP (1) | JP5160433B2 (ja) |
WO (1) | WO2007048033A1 (ja) |
Families Citing this family (89)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US20040152119A1 (en) * | 2003-02-05 | 2004-08-05 | Anup Sood | Solid phase sequencing |
EP1766075B1 (en) * | 2004-06-10 | 2010-03-17 | Amersham Biosciences Corp. | Method for nucleic acid analysis |
US7264934B2 (en) * | 2004-06-10 | 2007-09-04 | Ge Healthcare Bio-Sciences Corp. | Rapid parallel nucleic acid analysis |
US7692219B1 (en) | 2004-06-25 | 2010-04-06 | University Of Hawaii | Ultrasensitive biosensors |
US7563574B2 (en) * | 2006-03-31 | 2009-07-21 | Pacific Biosciences Of California, Inc. | Methods, systems and compositions for monitoring enzyme activity and applications thereof |
US9114398B2 (en) | 2006-11-29 | 2015-08-25 | Canon U.S. Life Sciences, Inc. | Device and method for digital multiplex PCR assays |
US8349167B2 (en) | 2006-12-14 | 2013-01-08 | Life Technologies Corporation | Methods and apparatus for detecting molecular interactions using FET arrays |
CA2672315A1 (en) * | 2006-12-14 | 2008-06-26 | Ion Torrent Systems Incorporated | Methods and apparatus for measuring analytes using large scale fet arrays |
US11339430B2 (en) | 2007-07-10 | 2022-05-24 | Life Technologies Corporation | Methods and apparatus for measuring analytes using large scale FET arrays |
US8262900B2 (en) * | 2006-12-14 | 2012-09-11 | Life Technologies Corporation | Methods and apparatus for measuring analytes using large scale FET arrays |
JP2010537643A (ja) | 2007-08-29 | 2010-12-09 | アプライド バイオシステムズ, エルエルシー | 代替的な核酸配列決定法 |
US8652781B2 (en) | 2008-02-12 | 2014-02-18 | Pacific Biosciences Of California, Inc. | Cognate sampling kinetics |
EP2274446B1 (en) | 2008-03-31 | 2015-09-09 | Pacific Biosciences of California, Inc. | Two slow-step polymerase enzyme systems and methods |
US8420366B2 (en) * | 2008-03-31 | 2013-04-16 | Pacific Biosciences Of California, Inc. | Generation of modified polymerases for improved accuracy in single molecule sequencing |
US8999676B2 (en) | 2008-03-31 | 2015-04-07 | Pacific Biosciences Of California, Inc. | Recombinant polymerases for improved single molecule sequencing |
EP2942404B1 (en) * | 2008-03-31 | 2016-11-23 | Pacific Biosciences of California, Inc. | Generation of modified polymerases for improved accuracy in single molecule sequencing |
WO2010008480A2 (en) | 2008-06-25 | 2010-01-21 | Ion Torrent Systems Incorporated | Methods and apparatus for measuring analytes using large scale fet arrays |
EP2307540B1 (en) | 2008-07-07 | 2017-04-19 | Oxford Nanopore Technologies Limited | Enzyme-pore constructs |
US20100036110A1 (en) * | 2008-08-08 | 2010-02-11 | Xiaoliang Sunney Xie | Methods and compositions for continuous single-molecule nucleic acid sequencing by synthesis with fluorogenic nucleotides |
US20100227327A1 (en) * | 2008-08-08 | 2010-09-09 | Xiaoliang Sunney Xie | Methods and compositions for continuous single-molecule nucleic acid sequencing by synthesis with fluorogenic nucleotides |
EP2326733A2 (en) * | 2008-09-05 | 2011-06-01 | Pacific Biosciences of California, Inc. | Engineering polymerases and reaction conditions for modified incorporation properties |
US20100301398A1 (en) | 2009-05-29 | 2010-12-02 | Ion Torrent Systems Incorporated | Methods and apparatus for measuring analytes |
US20100137143A1 (en) | 2008-10-22 | 2010-06-03 | Ion Torrent Systems Incorporated | Methods and apparatus for measuring analytes |
US20110014612A1 (en) | 2009-03-27 | 2011-01-20 | Life Technologies Corporation | Polymerase compositions & methods |
US8673627B2 (en) | 2009-05-29 | 2014-03-18 | Life Technologies Corporation | Apparatus and methods for performing electrochemical reactions |
US8776573B2 (en) | 2009-05-29 | 2014-07-15 | Life Technologies Corporation | Methods and apparatus for measuring analytes |
US8574835B2 (en) | 2009-05-29 | 2013-11-05 | Life Technologies Corporation | Scaffolded nucleic acid polymer particles and methods of making and using |
US20120261274A1 (en) | 2009-05-29 | 2012-10-18 | Life Technologies Corporation | Methods and apparatus for measuring analytes |
US20100311144A1 (en) | 2009-06-05 | 2010-12-09 | Life Technologies Corporation | Mutant dna polymerases |
JP5618556B2 (ja) | 2010-01-28 | 2014-11-05 | 株式会社日立ハイテクノロジーズ | 核酸分析装置,核酸分析反応デバイス、および核酸分析用反応デバイス用基板 |
CN106932456B (zh) | 2010-06-30 | 2020-02-21 | 生命科技公司 | 用于测试isfet阵列的方法和装置 |
AU2011226767B1 (en) | 2010-06-30 | 2011-11-10 | Life Technologies Corporation | Ion-sensing charge-accumulation circuits and methods |
CN114019006A (zh) | 2010-06-30 | 2022-02-08 | 生命科技公司 | 阵列列积分器 |
US11307166B2 (en) | 2010-07-01 | 2022-04-19 | Life Technologies Corporation | Column ADC |
JP5876044B2 (ja) | 2010-07-03 | 2016-03-02 | ライフ テクノロジーズ コーポレーション | 低濃度ドープドレインを有する化学的感応性センサ |
WO2012009206A2 (en) | 2010-07-12 | 2012-01-19 | Pacific Biosciences Of California, Inc. | Sequencing reactions with alkali metal cations for pulse width control |
US9618475B2 (en) | 2010-09-15 | 2017-04-11 | Life Technologies Corporation | Methods and apparatus for measuring analytes |
CN105911126B (zh) | 2010-09-24 | 2018-12-18 | 生命科技公司 | 匹配的晶体管对电路 |
US9970984B2 (en) | 2011-12-01 | 2018-05-15 | Life Technologies Corporation | Method and apparatus for identifying defects in a chemical sensor array |
US8747748B2 (en) | 2012-01-19 | 2014-06-10 | Life Technologies Corporation | Chemical sensor with conductive cup-shaped sensor surface |
US8821798B2 (en) | 2012-01-19 | 2014-09-02 | Life Technologies Corporation | Titanium nitride as sensing layer for microwell structure |
ES2683707T3 (es) | 2012-05-02 | 2018-09-27 | Ibis Biosciences, Inc. | Secuenciación de ADN |
EP3789502A1 (en) | 2012-05-02 | 2021-03-10 | Ibis Biosciences, Inc. | Dna sequencing |
EP2844775B1 (en) | 2012-05-02 | 2018-07-18 | Ibis Biosciences, Inc. | Dna sequencing |
EP2662692A1 (en) | 2012-05-11 | 2013-11-13 | Alacris Theranostics GmbH | Electrochemical detection of polymerase reactions by specific metal-phosphate complex formation |
US8786331B2 (en) | 2012-05-29 | 2014-07-22 | Life Technologies Corporation | System for reducing noise in a chemical sensor array |
GB2557509A (en) | 2012-06-08 | 2018-06-20 | Pacific Biosciences California Inc | Modified base detection with nanopore sequencing |
US9725768B2 (en) | 2012-08-31 | 2017-08-08 | Biovest International, Inc. | Methods for producing high-fidelity autologous idiotype vaccines |
US9399766B2 (en) | 2012-10-01 | 2016-07-26 | Pacific Biosciences Of California, Inc. | Recombinant polymerases for incorporation of protein shield nucleotide analogs |
US9080968B2 (en) | 2013-01-04 | 2015-07-14 | Life Technologies Corporation | Methods and systems for point of use removal of sacrificial material |
US9841398B2 (en) | 2013-01-08 | 2017-12-12 | Life Technologies Corporation | Methods for manufacturing well structures for low-noise chemical sensors |
US8962366B2 (en) | 2013-01-28 | 2015-02-24 | Life Technologies Corporation | Self-aligned well structures for low-noise chemical sensors |
US8963216B2 (en) | 2013-03-13 | 2015-02-24 | Life Technologies Corporation | Chemical sensor with sidewall spacer sensor surface |
US8841217B1 (en) | 2013-03-13 | 2014-09-23 | Life Technologies Corporation | Chemical sensor with protruded sensor surface |
US9835585B2 (en) | 2013-03-15 | 2017-12-05 | Life Technologies Corporation | Chemical sensor with protruded sensor surface |
US20140264471A1 (en) | 2013-03-15 | 2014-09-18 | Life Technologies Corporation | Chemical device with thin conductive element |
EP2972279B1 (en) | 2013-03-15 | 2021-10-06 | Life Technologies Corporation | Chemical sensors with consistent sensor surface areas |
US9116117B2 (en) | 2013-03-15 | 2015-08-25 | Life Technologies Corporation | Chemical sensor with sidewall sensor surface |
US20140264472A1 (en) | 2013-03-15 | 2014-09-18 | Life Technologies Corporation | Chemical sensor with consistent sensor surface areas |
US20140336063A1 (en) | 2013-05-09 | 2014-11-13 | Life Technologies Corporation | Windowed Sequencing |
US10458942B2 (en) | 2013-06-10 | 2019-10-29 | Life Technologies Corporation | Chemical sensor array having multiple sensors per well |
EP4354131A2 (en) | 2014-12-18 | 2024-04-17 | Life Technologies Corporation | High data rate integrated circuit with transmitter configuration |
US10077472B2 (en) | 2014-12-18 | 2018-09-18 | Life Technologies Corporation | High data rate integrated circuit with power management |
WO2016100521A1 (en) | 2014-12-18 | 2016-06-23 | Life Technologies Corporation | Methods and apparatus for measuring analytes using large scale fet arrays |
US10077470B2 (en) | 2015-07-21 | 2018-09-18 | Omniome, Inc. | Nucleic acid sequencing methods and systems |
US11155864B2 (en) * | 2015-07-30 | 2021-10-26 | Illumina, Inc. | Orthogonal deblocking of nucleotides |
WO2017087724A1 (en) | 2015-11-17 | 2017-05-26 | Omniome, Inc. | Methods for determining sequence profiles |
CN109072297B (zh) * | 2016-04-22 | 2022-09-13 | 加利福尼亚太平洋生物科学股份有限公司 | 使用增强的核苷酸特异性三元复合物形成检测的核酸测序方法和系统 |
SG11201809522WA (en) | 2016-04-29 | 2018-11-29 | Omniome Inc | Method of nucleic acid sequence determination |
AU2017258619B2 (en) | 2016-04-29 | 2020-05-14 | Pacific Biosciences Of California, Inc. | Sequencing method employing ternary complex destabilization to identify cognate nucleotides |
US10597643B2 (en) | 2016-04-29 | 2020-03-24 | Omniome, Inc. | Polymerases engineered to reduce nucleotide-independent DNA binding |
WO2018035134A1 (en) | 2016-08-15 | 2018-02-22 | Omniome, Inc. | Method and system for sequencing nucleic acids |
WO2018034780A1 (en) * | 2016-08-15 | 2018-02-22 | Omniome, Inc. | Sequencing method for rapid identification and processing of cognate nucleotide pairs |
CA3048415C (en) | 2016-12-30 | 2023-02-28 | Omniome, Inc. | Method and system employing distinguishable polymerases for detecting ternary complexes and identifying cognate nucleotides |
US9932631B1 (en) | 2017-09-11 | 2018-04-03 | Omniome, Inc. | Genotyping by polymerase binding |
WO2018136117A1 (en) * | 2017-01-20 | 2018-07-26 | Omniome, Inc. | Allele-specific capture of nucleic acids |
WO2018136487A1 (en) | 2017-01-20 | 2018-07-26 | Omniome, Inc. | Process for cognate nucleotide detection in a nucleic acid sequencing workflow |
US10161003B2 (en) | 2017-04-25 | 2018-12-25 | Omniome, Inc. | Methods and apparatus that increase sequencing-by-binding efficiency |
US9951385B1 (en) | 2017-04-25 | 2018-04-24 | Omniome, Inc. | Methods and apparatus that increase sequencing-by-binding efficiency |
CA3079411C (en) | 2017-10-19 | 2023-12-05 | Omniome, Inc. | Simultaneous background reduction and complex stabilization in binding assay workflows |
US20210040554A1 (en) * | 2018-03-13 | 2021-02-11 | Innovasion Labs, Inc. | Methods for single molecule sequencing |
AU2019276719A1 (en) | 2018-05-31 | 2020-11-26 | Pacific Biosciences Of California, Inc. | Increased signal to noise in nucleic acid sequencing |
US11180794B2 (en) | 2018-05-31 | 2021-11-23 | Omniome, Inc. | Methods and compositions for capping nucleic acids |
EP3608422A1 (en) | 2018-08-08 | 2020-02-12 | Amaryllis Innovation GmbH | Electrochemical detection of polymerase reactions by specific metal-phosphate complex formation |
US10768173B1 (en) | 2019-09-06 | 2020-09-08 | Element Biosciences, Inc. | Multivalent binding composition for nucleic acid analysis |
US10876148B2 (en) | 2018-11-14 | 2020-12-29 | Element Biosciences, Inc. | De novo surface preparation and uses thereof |
US10704094B1 (en) | 2018-11-14 | 2020-07-07 | Element Biosciences, Inc. | Multipart reagents having increased avidity for polymerase binding |
CN112368077B (zh) | 2018-12-07 | 2023-11-24 | 元素生物科学公司 | 流动池装置及其用途 |
US11287422B2 (en) | 2019-09-23 | 2022-03-29 | Element Biosciences, Inc. | Multivalent binding composition for nucleic acid analysis |
Family Cites Families (11)
Publication number | Priority date | Publication date | Assignee | Title |
---|---|---|---|---|
US4971903A (en) * | 1988-03-25 | 1990-11-20 | Edward Hyman | Pyrophosphate-based method and apparatus for sequencing nucleic acids |
US5516633A (en) * | 1991-08-15 | 1996-05-14 | Amersham Life Science, Inc. | DNA sequencing with a T7-type gene 6 exonuclease |
US5674679A (en) * | 1991-09-27 | 1997-10-07 | Amersham Life Science, Inc. | DNA cycle sequencing |
US5432065A (en) * | 1993-03-30 | 1995-07-11 | United States Biochemical Corporation | Cycle sequencing with non-thermostable DNA polymerases |
US20030108867A1 (en) * | 1999-04-20 | 2003-06-12 | Chee Mark S | Nucleic acid sequencing using microsphere arrays |
EP1182267B1 (en) * | 2000-03-30 | 2012-01-18 | Toyota Jidosha Kabushiki Kaisha | Method of determining base sequence of single nucleic acid molecule |
GB0103622D0 (en) * | 2001-02-14 | 2001-03-28 | Univ Cambridge Tech | Methods for detecting DNA polymerisation |
CA2440754A1 (en) * | 2001-03-12 | 2002-09-19 | Stephen Quake | Methods and apparatus for analyzing polynucleotide sequences by asynchronous base extension |
JP2007525151A (ja) * | 2003-01-29 | 2007-09-06 | 454 コーポレーション | 一本鎖dnaライブラリーの調製方法 |
EP1766075B1 (en) * | 2004-06-10 | 2010-03-17 | Amersham Biosciences Corp. | Method for nucleic acid analysis |
US7264934B2 (en) * | 2004-06-10 | 2007-09-04 | Ge Healthcare Bio-Sciences Corp. | Rapid parallel nucleic acid analysis |
-
2005
- 2005-10-21 US US11/255,683 patent/US7264934B2/en active Active
-
2006
- 2006-10-20 WO PCT/US2006/041232 patent/WO2007048033A1/en active Application Filing
- 2006-10-20 EP EP06826449.8A patent/EP1954827B1/en active Active
- 2006-10-20 JP JP2008536859A patent/JP5160433B2/ja active Active
Also Published As
Publication number | Publication date |
---|---|
US7264934B2 (en) | 2007-09-04 |
EP1954827B1 (en) | 2013-07-24 |
WO2007048033A1 (en) | 2007-04-26 |
EP1954827A1 (en) | 2008-08-13 |
JP2009512452A (ja) | 2009-03-26 |
EP1954827A4 (en) | 2009-12-09 |
US20060051807A1 (en) | 2006-03-09 |
Similar Documents
Publication | Publication Date | Title |
---|---|---|
JP5160433B2 (ja) | 迅速並行核酸分析 | |
JP5037343B2 (ja) | 核酸分析方法 | |
RU2698125C2 (ru) | Библиотеки для секвенирования нового поколения | |
US9758825B2 (en) | Centroid markers for image analysis of high density clusters in complex polynucleotide sequencing | |
EP1937702A2 (en) | Labeled nucleotide analogs and uses therefor | |
JP2010515433A (ja) | ライゲーション増幅 | |
JP2006513705A (ja) | 核酸増殖 | |
JP2018518947A (ja) | ヌクレオチドのオルトゴナルな脱ブロッキング | |
JP2002531106A (ja) | 不連続プライマ−伸長による核酸反復配列の長さ決定 | |
WO2020093261A1 (zh) | 对多核苷酸进行测序的方法 | |
JP4297966B2 (ja) | 標識ヌクレオチドを利用することによる核酸の新規測定方法 | |
US7838236B2 (en) | Analytical method and kit | |
US9777319B2 (en) | Method for isothermal DNA amplification starting from an RNA template | |
KR20030092136A (ko) | 분석 방법 및 키트 | |
EP3625240A1 (en) | Sequencing using non-natural nucleotides | |
US20230348969A1 (en) | Methods and systems for nucleic acid sequencing |
Legal Events
Date | Code | Title | Description |
---|---|---|---|
A621 | Written request for application examination |
Free format text: JAPANESE INTERMEDIATE CODE: A621 Effective date: 20091016 |
|
RD02 | Notification of acceptance of power of attorney |
Free format text: JAPANESE INTERMEDIATE CODE: A7422 Effective date: 20091016 |
|
RD04 | Notification of resignation of power of attorney |
Free format text: JAPANESE INTERMEDIATE CODE: A7424 Effective date: 20091016 |
|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20101001 |
|
A131 | Notification of reasons for refusal |
Free format text: JAPANESE INTERMEDIATE CODE: A131 Effective date: 20120424 |
|
A601 | Written request for extension of time |
Free format text: JAPANESE INTERMEDIATE CODE: A601 Effective date: 20120720 |
|
A602 | Written permission of extension of time |
Free format text: JAPANESE INTERMEDIATE CODE: A602 Effective date: 20120727 |
|
A521 | Request for written amendment filed |
Free format text: JAPANESE INTERMEDIATE CODE: A523 Effective date: 20121022 |
|
TRDD | Decision of grant or rejection written | ||
A01 | Written decision to grant a patent or to grant a registration (utility model) |
Free format text: JAPANESE INTERMEDIATE CODE: A01 Effective date: 20121113 |
|
A61 | First payment of annual fees (during grant procedure) |
Free format text: JAPANESE INTERMEDIATE CODE: A61 Effective date: 20121212 |
|
R150 | Certificate of patent or registration of utility model |
Ref document number: 5160433 Country of ref document: JP Free format text: JAPANESE INTERMEDIATE CODE: R150 Free format text: JAPANESE INTERMEDIATE CODE: R150 |
|
FPAY | Renewal fee payment (event date is renewal date of database) |
Free format text: PAYMENT UNTIL: 20151221 Year of fee payment: 3 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
S531 | Written request for registration of change of domicile |
Free format text: JAPANESE INTERMEDIATE CODE: R313531 |
|
S533 | Written request for registration of change of name |
Free format text: JAPANESE INTERMEDIATE CODE: R313533 |
|
R350 | Written notification of registration of transfer |
Free format text: JAPANESE INTERMEDIATE CODE: R350 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |
|
R250 | Receipt of annual fees |
Free format text: JAPANESE INTERMEDIATE CODE: R250 |