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    Reagents, methods, and libraries for bead-based sequencing
    5.
    发明申请
    Reagents, methods, and libraries for bead-based sequencing 审中-公开

    公开(公告)号：US20080003571A1

    公开(公告)日：2008-01-03

    申请号：US11345979

    申请日：2006-02-01

    申请人： Kevin McKernan ,  Alan Blanchard ,  Lev Kotler ,  Gina Costa

    发明人： Kevin McKernan ,  Alan Blanchard ,  Lev Kotler ,  Gina Costa

    IPC分类号： C12Q1/68

    CPC分类号： C12Q1/6874 ,  B82Y15/00 ,  B82Y30/00 ,  C12Q1/68 ,  C12Q1/6837 ,  C12Q1/6844 ,  C12Q1/6869 ,  C12Q2565/537 ,  C12Q2565/518 ,  C12Q2565/513 ,  C12Q2565/137 ,  C12Q2565/102 ,  C12Q2533/107 ,  C12Q2537/165

    摘要： The present invention provides methods for determining a nucleic acid sequence by performing successive cycles of duplex extension along a single stranded template. The cycles comprise steps of extension, ligation, and, preferably, cleavage. In certain embodiments the methods make use of extension probes containing phosphorothiolate linkages and employ agents appropriate to cleave such linkages. In certain embodiments the methods make use of extension probes containing an abasic residue or a damaged base and employ agents appropriate to cleave linkages between a nucleoside and an abasic residue and/or agents appropriate to remove a damaged base from a nucleic acid. The invention provides methods of determining information about a sequence using at least two distinguishably labeled probe families. In certain embodiments the methods acquire less than 2 bits of information from each of a plurality of nucleotides in the template in each cycle. In certain embodiments the sequencing reactions are performed on templates attached to beads, which are immobilized in or on a semi-solid support. The invention further provides sets of labeled extension probes containing phosphorothiolate linkages or trigger residues that are suitable for use in the method. In addition, the invention includes performing multiple sequencing reactions on a single template by removing initializing oligonucleotides and extended strands and performing subsequent reactions using different initializing oligonucleotides. The invention further provides efficient methods for preparing templates, particularly for performing sequencing multiple different templates in parallel. The invention also provides methods for performing ligation and cleavage. The invention also provides new libraries of nucleic acid fragments containing paired tags, and methods of preparing microparticles having multiple different templates (e.g., containing paired tags) attached thereto and of sequencing the templates individually. The invention also provides automated sequencing systems, flow cells, image processing methods, and computer-readable media that store computer-executable instructions (e.g., to perform the image-processing methods) and/or sequence information. In certain embodiments the sequence information is stored in a database.

    Reagents, methods, and libraries for bead-based sequencing
    6.
    发明授权
    Reagents, methods, and libraries for bead-based sequencing 有权
    用于珠基测序的试剂，方法和文库

    公开(公告)号：US08329404B2

    公开(公告)日：2012-12-11

    申请号：US13410919

    申请日：2012-03-02

    申请人： Kevin McKernan ,  Alan Blanchard ,  Lev Kotler ,  Gina Costa

    发明人： Kevin McKernan ,  Alan Blanchard ,  Lev Kotler ,  Gina Costa

    IPC分类号： C12Q1/68

    CPC分类号： C12Q1/6874 ,  B82Y15/00 ,  B82Y30/00 ,  C12Q1/68 ,  C12Q1/6837 ,  C12Q1/6844 ,  C12Q1/6869 ,  C12Q2565/537 ,  C12Q2565/518 ,  C12Q2565/513 ,  C12Q2565/137 ,  C12Q2565/102 ,  C12Q2533/107 ,  C12Q2537/165

    摘要： The present invention provides methods for determining a nucleic acid sequence by performing successive cycles of duplex extension along a single stranded template. The cycles comprise steps of extension, ligation, and, preferably, cleavage. In certain embodiments the methods make use of extension probes containing phosphorothiolate linkages and employ agents appropriate to cleave such linkages. The invention provides methods of determining information about a sequence using at least two distinguishably labeled probe families. In certain embodiments the methods acquire less than 2 bits of information from each of a plurality of nucleotides in the template in each cycle. In certain embodiments the sequencing reactions are performed on templates attached to immobilized beads. The invention further provides sets of labeled extension probes containing phosphorothiolate linkages. In addition, the invention includes performing multiple sequencing reactions on a single template by removing initializing oligonucleotides and extended strands and performing subsequent reactions using different initializing oligonucleotides.

    摘要翻译： 本发明提供了通过沿着单链模板进行连续循环的双链延伸来确定核酸序列的方法。 循环包括延伸，连接和优选切割的步骤。 在某些实施方案中，所述方法利用含有硫代磷酸酯键的延伸探针，并使用适合切割这种连接的试剂。 本发明提供使用至少两个可区分标记的探针家族确定关于序列的信息的方法。 在某些实施方案中，该方法在每个周期中从模板中的多个核苷酸中的每一个获取少于2位的信息。 在某些实施方案中，测序反应在附着于固定化珠粒的模板上进行。 本发明还提供了含有硫代磷酸酯键的标记的延伸探针的集合。 此外，本发明包括通过除去初始化寡核苷酸和延伸的链并使用不同的初始化寡核苷酸进行后续反应，在单个模板上进行多个测序反应。

    Reagents, Methods, and Libraries for Bead-Based Sequencing
    7.
    发明申请
    Reagents, Methods, and Libraries for Bead-Based Sequencing 审中-公开

    公开(公告)号：US20100297626A1

    公开(公告)日：2010-11-25

    申请号：US12628209

    申请日：2009-11-30

    申请人： Kevin MCKERNAN ,  Alan Blanchard ,  Lev Kotler ,  Gina Costa

    发明人： Kevin MCKERNAN ,  Alan Blanchard ,  Lev Kotler ,  Gina Costa

    IPC分类号： C12Q1/68

    CPC分类号： C12Q1/6874 ,  B82Y15/00 ,  B82Y30/00 ,  C12Q1/68 ,  C12Q1/6837 ,  C12Q1/6844 ,  C12Q1/6869 ,  C12Q2565/537 ,  C12Q2565/518 ,  C12Q2565/513 ,  C12Q2565/137 ,  C12Q2565/102 ,  C12Q2533/107 ,  C12Q2537/165

    摘要： The present invention provides methods for determining a nucleic acid sequence by performing successive cycles of duplex extension along a single stranded template. The cycles comprise steps of extension, ligation, and, preferably, cleavage. In certain embodiments the methods make use of extension probes containing phosphorothiolate linkages and employ agents appropriate to cleave such linkages. In certain embodiments the methods make use of extension probes containing an abasic residue or a damaged base and employ agents appropriate to cleave linkages between a nucleoside and an abasic residue and/or agents appropriate to remove a damaged base from a nucleic acid. The invention provides methods of determining information about a sequence using at least two distinguishably labeled probe families. In certain embodiments the methods acquire less than 2 bits of information from each of a plurality of nucleotides in the template in each cycle. In certain embodiments the sequencing reactions are performed on templates attached to beads, which are immobilized in or on a semi-solid support. The invention further provides sets of labeled extension probes containing phosphorothiolate linkages or trigger residues that are suitable for use in the method. In addition, the invention includes performing multiple sequencing reactions on a single template by removing initializing oligonucleotides and extended strands and performing subsequent reactions using different initializing oligonucleotides. The invention further provides efficient methods for preparing templates, particularly for performing sequencing multiple different templates in parallel. The invention also provides methods for performing ligation and cleavage. The invention also provides new libraries of nucleic acid fragments containing paired tags, and methods of preparing microparticles having multiple different templates (e.g., containing paired tags) attached thereto and of sequencing the templates individually. The invention also provides automated sequencing systems, flow cells, image processing methods, and computer-readable media that store computer-executable instructions (e.g., to perform the image-processing methods) and/or sequence information. In certain embodiments the sequence information is stored in a database.

    Reagents, methods, and libraries for bead-based sequencing
    8.
    发明申请
    Reagents, methods, and libraries for bead-based sequencing 有权
    用于珠基测序的试剂，方法和文库

    公开(公告)号：US20090181860A1

    公开(公告)日：2009-07-16

    申请号：US12220208

    申请日：2008-07-21

    申请人： Kevin McKernan ,  Alan Blanchard ,  Lev Kotler ,  Gina Costa

    发明人： Kevin McKernan ,  Alan Blanchard ,  Lev Kotler ,  Gina Costa

    IPC分类号： C40B40/06 ,  C07H21/04

    CPC分类号： C12Q1/6874 ,  B82Y15/00 ,  B82Y30/00 ,  C12Q1/68 ,  C12Q1/6837 ,  C12Q1/6844 ,  C12Q1/6869 ,  C12Q2565/537 ,  C12Q2565/518 ,  C12Q2565/513 ,  C12Q2565/137 ,  C12Q2565/102 ,  C12Q2533/107 ,  C12Q2537/165

    摘要： The present invention provides methods for determining a nucleic acid sequence by performing successive cycles of duplex extension along a single stranded template. The cycles comprise steps of extension, ligation, and, preferably, cleavage. In certain embodiments the methods make use of extension probes containing phosphorothiolate linkages and employ agents appropriate to cleave such linkages. The invention provides methods of determining information about a sequence using at least two distinguishably labeled probe families. In certain embodiments the methods acquire less than 2 bits of information from each of a plurality of nucleotides in the template in each cycle. In certain embodiments the sequencing reactions are performed on templates attached to immobilized beads. The invention further provides sets of labeled probes containing phosphorothiolate linkages. In addition, the invention includes performing multiple sequencing reactions on a single template by removing initializing oligonucleotides and extended strands and performing subsequent reactions using different initializing oligonucleotides.

    摘要翻译： 本发明提供了通过沿着单链模板进行连续循环的双链延伸来确定核酸序列的方法。 循环包括延伸，连接和优选切割的步骤。 在某些实施方案中，所述方法利用含有硫代磷酸酯键的延伸探针，并使用适合切割这种连接的试剂。 本发明提供使用至少两个可区分标记的探针家族确定关于序列的信息的方法。 在某些实施方案中，该方法在每个周期中从模板中的多个核苷酸中的每一个获取少于2位的信息。 在某些实施方案中，测序反应在附着于固定化珠粒的模板上进行。 本发明还提供了含有硫代磷酸酯键的标记探针的集合。 此外，本发明包括通过除去初始化寡核苷酸和延伸的链并使用不同的初始化寡核苷酸进行后续反应，在单个模板上进行多个测序反应。