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1. 发明申请

WO2004072297A3 TERMINAL-PHOSPHATE-LABELED NUCLEOTIDES AND METHODS OF USE 审中-公开
标题翻译：终端磷酸酯标记的核苷酸和使用方法
公开(公告)号：WO2004072297A3
公开(公告)日：2007-01-18
申请号：PCT/US2004002785
申请日：2004-01-30
申请人： AMERSHAM BIOSCIENCES CORP , FULLER CARL , KUMAR SHIV , SOOD ANUP , NELSON JOHN
发明人： FULLER CARL , KUMAR SHIV , SOOD ANUP , NELSON JOHN
IPC分类号： C12Q1/68 , C07H19/10 , C07H19/20 , C07H21/00 , C12Q20060101 , C12Q1/00 , C12Q1/42 , G01N33/52 , G01N33/58
CPC分类号： C12Q1/6823 , C07H19/10 , C07H19/20 , C07H21/00 , C12Q1/6816 , C12Q1/6851 , C12Q2521/525
摘要： The present invention relates to improved methods of detecting a target using a labeled substrate or substrate analog. The methods comprise reacting the substrate or substrate analog in an enzyme-catalyzed reaction which produces a labeled moiety with independently detectable signal only when such substrate or substrate analog reacts. The present invention, in particular, describes methods of detecting a nucleic acid in a sample, based on the use of terminal-phosphate-labeled nucleotides as substrate for nucleic acid polymerases. The methods provided by this invention utilize a nucleoside polyphosphate, dideoxynucleoside polyphosphate, or deoxynucleoside polyphosphate analogue which has a colorimetric dye, chemiluminescent, or fluorescent moiety, a mass tag or an electrochemical tag attached to the terminal-phosphate. When a nucleic acid polymerase uses this analogue as a substrate, an enzyme-activatable label would be present on the inorganic polyphosphate by-product of phosphoryl transfer. Cleavage of the polyphosphate product of phosphoryl transfer via phosphatase leads to a detectable change in the label attached thereon. When the polymerase assay is performed in the presence of a phosphatase, there is provided a convenient method for real-time monitoring of DNA or RNA synthesis and detection of a target nucleic acid.
摘要翻译：本发明涉及使用标记的底物或底物类似物检测靶的改进方法。所述方法包括在酶催化的反应中使底物或底物类似物反应，其仅在这种底物或底物类似物反应时产生具有独立可检测信号的标记部分。本发明特别地描述了基于使用末端磷酸酯标记的核苷酸作为核酸聚合酶的底物来检测样品中核酸的方法。本发明提供的方法利用了具有比色染料，化学发光或荧光部分的核苷多磷酸，双脱氧核苷多聚磷酸酯或脱氧核苷多聚磷酸酯类似物，连接至末端磷酸酯的质量标签或电化学标签。当核酸聚合酶使用该类似物作为底物时，酶活性标记将存在于磷酸转移的无机多磷酸盐副产物上。通过磷酸酶切割磷酸转移的多磷酸盐产物导致附着在其上的标记物的可检测的变化。当在磷酸酶的存在下进行聚合酶测定时，提供了用于实时监测DNA或RNA合成和检测靶核酸的方便的方法。

2. 发明申请

WO2004071155A2 SOLID PHASE SEQUENCING 审中-公开
标题翻译：固相序列
公开(公告)号：WO2004071155A2
公开(公告)日：2004-08-26
申请号：PCT/US2004003283
申请日：2004-02-05
申请人： AMERSHAM BIOSCIENCES CORP , SOOD ANUP , KUMAR SHIV , NELSON JOHN , FULLER CARL
发明人： SOOD ANUP , KUMAR SHIV , NELSON JOHN , FULLER CARL
IPC分类号： C12P19/34 , C12Q1/68 , G01R29/18
CPC分类号： C12Q1/6858 , C12Q1/6874 , C12Q2565/537 , C12Q2535/125 , C12Q2521/525 , C12Q2525/101
摘要： The present invention describes methods of sequencing a nucleic acid in a sample, based on the use of terminal-phosphate-labeled nucleotides as substrates for nucleic acid polymerases. The methods provided by this invention utilize a nucleoside polyphosphate, dideoxynucleoside polyphosphate, or deoxynucleoside polyphosphate analogue which has a colorimetric dye, chemiluminescent, or fluorescent moiety, a mass tag or an electrochemical tag attached to the terminal-phosphate. When a nucleic acid polymerase uses this analogue as a substrate, an enzyme-activatable label would be present on the inorganic polyphosphate by-product of phosphoryl transfer. Cleavage of the polyphosphate product of phosphoryl transfer via phosphatase leads to a detectable change in the label attached thereon. In some instances the labeled polyphosphate may be detected directly via the label and provide information on the nucleic acid. When the polymerase assay is performed in the presence of a phosphatase, there is provided a convenient method for real-time monitoring of DNA or RNA synthesis and characterization of a target nucleic acid.
摘要翻译：本发明描述了基于使用末端磷酸酯标记的核苷酸作为核酸聚合酶的底物来测序样品中核酸的方法。本发明提供的方法利用具有比色染料，化学发光或荧光部分的核苷多磷酸，双脱氧核苷多磷酸酯或脱氧核苷多聚磷酸酯类似物，连接至末端磷酸酯的质量标签或电化学标签。当核酸聚合酶使用该类似物作为底物时，酶活性标记将存在于磷酸转移的无机多磷酸盐副产物上。通过磷酸酶切割磷酸转移的多磷酸盐产物导致附着在其上的标记物的可检测的变化。在一些情况下，标记的多磷酸盐可以通过标记物直接检测并提供关于核酸的信息。当在磷酸酶存在下进行聚合酶测定时，提供了用于实时监测DNA或RNA合成和靶核酸表征的方便的方法。

3. 发明申请

WO2004072238A3 TERMINAL-PHOSPHATE-LABELED NUCLEOTIDES WITH NEW LINKERS 审中-公开
标题翻译：带有新连接器的终端磷酸酯标记的核苷酸
公开(公告)号：WO2004072238A3
公开(公告)日：2006-05-18
申请号：PCT/US2004003284
申请日：2004-02-05
申请人： AMERSHAM BIOSCIENCES CORP , KUMAR SHIV , MCDOUGALL MARK , SOOD ANUP , NELSON JOHN , FULLER CARL , MACKLIN JOHN , MITSIS PAUL
发明人： KUMAR SHIV , MCDOUGALL MARK , SOOD ANUP , NELSON JOHN , FULLER CARL , MACKLIN JOHN , MITSIS PAUL
IPC分类号： C12Q1/68 , C12N20060101 , C12N1/00 , C12N9/00 , C12N9/22 , C12P19/34
CPC分类号： C07H19/10 , C07H19/20 , C07H21/00 , C12P19/34
摘要： The present invention describes methods of using terminal-phosphate-labeled nucleotides in the presence of a manganese salt to enhance their substrate properties towards various enzymes. Particularly described are methods of detecting a nucleic acid in a sample, based on the use of terminal-phosphate-labeled nucleotides as substrates for nucleic acid polymerases, in the presence of a manganese salt. Further provided are manganese complexes of terminal-phosphate-labeled nucleotides as well as terminal-phosphate-labeled nucleotides with new linkers with enhanced substrate properties.
摘要翻译：本发明描述了在锰盐存在下使用末端磷酸酯标记的核苷酸以增强其对各种酶的底物性质的方法。特别描述的是在锰盐存在的情况下，基于使用末端磷酸酯标记的核苷酸作为核酸聚合酶的底物，检测样品中的核酸的方法。还提供了末端磷酸酯标记的核苷酸的锰络合物以及具有增强的底物性质的新接头的末端磷酸酯标记的核苷酸。

4. 发明申请

WO2004085539A3 CYANINE DYE LABELLING REAGENTS WITH MESO-SUBSTITUTION 审中-公开
标题翻译：具有MESO替代品的氰基染料标签试剂
公开(公告)号：WO2004085539A3
公开(公告)日：2006-01-12
申请号：PCT/US2004007833
申请日：2004-03-12
申请人： AMERSHAM BIOSCIENCES CORP , CHEN CHUNG-YUAN , KUMAR SHIV
发明人： CHEN CHUNG-YUAN , KUMAR SHIV
IPC分类号： C07D209/24 , C09B23/01 , C09B23/02 , G01N33/58 , G01N33/533 , C07D209/02 , C07D263/62 , C07D277/60 , C07K17/06 , G01N33/566
CPC分类号： C09B23/164 , C07D209/24 , C09B23/0058 , C09B23/02 , G01N33/582
摘要： Disclosed are cyanine dyes having meso-substituents in the polymethine chain linking the heterocyclic ring systems. The dyes are of formula (I) in which groups R
摘要翻译：公开了在连接杂环体系的聚甲炔链中具有内消旋取代基的花青染料。染料具有式（I），其中R 1〜

5. 发明申请

WO2004029579A3 FLUORESCENT LABELING REAGENTS WITH MULTIPLE DONORS AND ACCEPTORS 审中-公开
标题翻译：荧光标签试剂与多个捐助者和接受者
公开(公告)号：WO2004029579A3
公开(公告)日：2004-08-19
申请号：PCT/US0330361
申请日：2003-09-25
申请人： AMERSHAM BIOSCIENCES CORP , KUMAR SHIV , CHEN CHUNG-YUAN
发明人： KUMAR SHIV , CHEN CHUNG-YUAN
IPC分类号： C07D311/82 , C07D405/14 , C07D473/02 , C07D487/04 , C07D491/22 , C09B11/04 , C09B11/08 , C09B11/24 , C12Q1/68 , C07D311/80
CPC分类号： C09B11/08 , C07D311/82 , C07D405/14 , C07D487/04 , C09B11/04 , C09B11/24 , C12Q1/6869 , C12Q2527/125
摘要： Fluorescent labeling reagents with multiple donors and acceptors, based on and synthesized from easily prepared dye building blocks.
摘要翻译：荧光标记试剂与多个供体和受体，基于并由易于制备的染料积木合成。

6. 发明申请

WO2004029578A3 ENERGY TRANSFER DYES, TERMINATORS AND USE THEREOF 审中-公开
标题翻译：能量转移染料，终止剂及其用途
公开(公告)号：WO2004029578A3
公开(公告)日：2004-07-08
申请号：PCT/US0330360
申请日：2003-09-25
申请人： AMERSHAM BIOSCIENCES CORP , KUMAR SHIV , CHEN CHUNG-YUAN , RAO SUDHAKAR
发明人： KUMAR SHIV , CHEN CHUNG-YUAN , RAO SUDHAKAR
IPC分类号： C07D311/82 , C07D405/14 , C07D473/02 , C07D487/04 , C07D491/22 , C09B11/04 , C09B11/08 , C09B11/24 , C12Q1/68 , C07H21/04 , C12P19/34
CPC分类号： C09B11/08 , C07D311/82 , C07D405/14 , C07D487/04 , C09B11/04 , C09B11/24 , C12Q1/6869 , C12Q2527/125
摘要： The present invention relates to a novel set of four fluorescently labeled dye terminators. Two of them are single dye labeled terminators and the other two dye terminators are based on fluorescent resonance energy transfer (FRET). The FRET dye terminators are generated from the 4', 5'-bis-aminomethylfluorescein. Of the two amino groups of the donor dye, 4', 5'-bis-aminomethylfluorescein, one amino group is used to attach the acceptor dye, and the other amino group is used to attach the dideoxynucleoside-5'-triphosphate. Their preparation, energy transfer efficiency, and use as labels, specifically, in DNA sequencing reactions is disclosed.
摘要翻译：本发明涉及一组新的四种荧光标记的染料终止剂。其中两个是单染料标记的终止子，另两个染料终止子基于荧光共振能量转移（FRET）。 FRET染料终止子由4'，5'-双 - 氨基甲基荧光素产生。在供体染料的两个氨基中，4'，5'-双氨基甲基荧光素，一个氨基用于连接受体染料，另一个氨基用于连接双脱氧核苷-5'-三磷酸。它们的制备，能量转移效率和作为标记的用途，特别是在DNA测序反应中被公开。

7. 发明申请

WO2004020604A3 ALLELE SPECIFIC PRIMER EXTENSION 审中-公开
标题翻译： ALLELE特定的PRIMER扩展
公开(公告)号：WO2004020604A3
公开(公告)日：2004-08-12
申请号：PCT/US0327286
申请日：2003-08-29
申请人： AMERSHAM BIOSCIENCES CORP
发明人： SOOD ANUP , KUMAR SHIV , FULLER CARL , NELSON JOHN , MACKLIN JOHN
IPC分类号： C12N15/09 , C12Q1/42 , C12Q1/44 , C12Q1/68 , C12P19/34
CPC分类号： C12Q1/6869 , C12Q1/6823 , C12Q2600/156 , C12Q2535/125 , C12Q2525/125 , C12Q2521/125 , C12Q2565/301 , C12Q2525/101
摘要： A method of characterizing a nucleic acid sample is provided that includes the steps of: (a) conducting a DNA polymerase reaction that includes the reaction of a template, an allele specific primer, at least one terminal phosphate-labeled nucleotide, DNA polymerase, and optionally an enzyme having 3' ? 5' exonuclease activity when the primer is non-hydrolyzable, which reaction results in the production of labeled polyphosphate; (b) permitting the labeled polyphosphate to react with a phosphatase to produce a detectable species; (c) detecting the detectable species; and (d) characterizing the nucleic acid sample based on such detection.
摘要翻译：提供了表征核酸样品的方法，其包括以下步骤：（a）进行DNA聚合酶反应，其包括模板，等位基因特异性引物，至少一个末端磷酸酯标记的核苷酸，DNA聚合酶和任选地具有3' 5'核酸外切酶活性，该反应导致标记多磷酸盐的产生; （b）允许标记的多磷酸酯与磷酸酶反应以产生可检测的物质; （c）检测可检测物种; 和（d）基于这种检测来表征核酸样品。

8. 发明申请

WO2004020602A3 TERMINAL PHOSPHATE BLOCKED NUCLEOSIDE POLYPHOSPHATES 审中-公开
标题翻译：磷酸盐封闭的核苷酸多磷酸盐
公开(公告)号：WO2004020602A3
公开(公告)日：2004-05-13
申请号：PCT/US0327284
申请日：2003-08-29
申请人： AMERSHAM BIOSCIENCES CORP
发明人： SOOD ANUP , KUMAR SHIV , FULLER CARL , NELSON JOHN
IPC分类号： C12N15/09 , C07H21/00 , C07H21/02 , C07H21/04 , C12N20060101 , C12P19/34 , C12Q1/68
CPC分类号： C12Q1/686 , C12Q2525/186 , C12Q2531/119
摘要： The present invention describes terminal phosphate blocked nucleoside polyphosphates that are stable at high temperature and their use in nucleic acid amplification and analysis. Current invention further describes charge modified terminal phosphate blocked nucleoside polyphosphates for improved incorporation and direct loading of nucleic acid sequencing reactions onto separating media.
摘要翻译：本发明描述了在高温下稳定的末端磷酸封端的核苷多磷酸盐及其在核酸扩增和分析中的用途。本发明进一步描述了电荷修饰的末端磷酸封端的核苷多磷酸酯，用于改善引入并将核酸测序反应直接加载到分离介质上。

9. 发明申请

WO2004020603A3 ANALYTE DETECTION 审中-公开
标题翻译：分析检测
公开(公告)号：WO2004020603A3
公开(公告)日：2004-04-22
申请号：PCT/US0327285
申请日：2003-08-29
申请人： AMERSHAM BIOSCIENCES CORP
发明人： SOOD ANUP , KUMAR SHIV , FULLER CARL , NELSON JOHN
IPC分类号： C12N15/09 , C12P19/34 , C12Q1/02 , C12Q1/42 , C12Q1/68
CPC分类号： C12Q1/68 , C12Q1/6823 , C12Q1/6844 , C12Q2565/301 , C12Q2563/179 , C12Q2525/301 , C12Q2521/525 , C12Q2521/101
摘要： A method of characterizing an analyte sample is provided that includes the steps of: (a) anchoring the analyte to a nucleic acid template of known sequence; (b) conducting a DNA polymerase reaction that includes the reaction of a template, a non-hydrolyzable primer, at least one terminal phosphate-labeled nucleotide, DNA polymerase, and an enzyme having 3' 5' exonuclease activity which reaction results in the production of labeled polyphosphate; (c) permitting the labeled polyphosphate to react with a phosphatase to produce a detectable species characteristic of the sample; (d) detecting the detectable species. The method may include the step of characterizing the nucleic acid sample based on the detection. Also provided are methods of analyzing multiple analytes in a sample, and kits for characterizing analyte samples.
摘要翻译：提供表征分析物样品的方法，其包括以下步骤：（a）将分析物锚定到已知序列的核酸模板; （b）进行DNA聚合酶反应，其包括模板，不可水解的引物，至少一个末端磷酸酯标记的核苷酸，DNA聚合酶和具有3'5'核酸外切酶活性的酶的反应，该反应导致生产的标记多磷酸盐; （c）允许标记的多磷酸酯与磷酸酶反应以产生样品的可检测物种特征; （d）检测可检测物种。该方法可以包括基于检测来表征核酸样品的步骤。还提供了分析样品中多种分析物的方法，以及用于表征分析物样品的试剂盒。

10. 发明申请

WO03020734A3 LABELED NUCLEOSIDE POLYPHOSPHATES 审中-公开
标题翻译：标记核苷酸多磷酸酯
公开(公告)号：WO03020734A3
公开(公告)日：2003-06-12
申请号：PCT/US0227565
申请日：2002-08-29
申请人： AMERSHAM BIOSCIENCES CORP
发明人： KUMAR SHIV , SOOD ANUP
IPC分类号： G01N27/62 , C07H19/10 , C07H19/20 , C07H21/00 , C12N15/09 , C12Q1/42 , C12Q1/68 , G01N21/76 , G01N21/78 , G01N33/543 , G01N33/58 , C07H19/00 , C07H21/02 , C07H21/04 , C12P19/34
CPC分类号： C07H19/10 , C07H19/20 , C07H21/00 , C12Q1/6851
摘要： The present invention describes new compositions of matter in the form of labeled nucleoside polyphosphates with four or more phosphates. In addition compositions of nucleoside polyphosphates with four or more phosphates that are substrates for nucleic acid polymerases with enhanced substrate properties and methods of using these nucleoside polyphosphates for nucleic acid detection, characterization and quantification are described. The compositions provided by this invention include nucleoside polyphosphate, dideoxynucleoside polyphosphate, or deoxynucleoside polyphosphate analogues which have colorimetric, chemiluminescent, or fluorescent moieties, mass tags or an electrochemical tags attached to the terminal-phosphate. When a nucleic acid polymerase uses this analogue as a substrate, an enzyme-activatable label would be present on the inorganic polyphosphate by-product of phosphoryl transfer. Cleavage of the polyphosphate product of phosphoryl transfer via phosphatase leads to a detectable change in the label attached thereon. When the polymerase assay is performed in the presence of a phosphatase, there is provided a convenient method for real-time monitoring of DNA or RNA synthesis and detection of a target nucleic acid.
摘要翻译：本发明描述了具有四种或更多种磷酸盐的标记的核苷多磷酸盐形式的物质的新组合物。此外，还描述了具有四种或更多种磷酸盐的核苷多磷酸盐的组合物，其是具有增强的底物性质的核酸聚合酶的底物和使用这些核苷多磷酸盐用于核酸检测，表征和定量的方法。本发明提供的组合物包括具有比色，化学发光或荧光部分的核苷多磷酸，双脱氧核苷多磷酸或脱氧核苷多磷酸酯类似物，质粒标签或连接至末端磷酸的电化学标记。当核酸聚合酶使用该类似物作为底物时，酶活性标记将存在于磷酸转移的无机多磷酸盐副产物上。通过磷酸酶切割磷酸转移的多磷酸盐产物导致附着在其上的标记物的可检测的变化。当在磷酸酶的存在下进行聚合酶测定时，提供了用于实时监测DNA或RNA合成和检测靶核酸的方便的方法。

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