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1. 发明公开

KR1020070112785A 비드-기초 서열화를 위한 시약, 방법, 및 라이브러리 无效
标题翻译：用于基于BEAD的测序的试剂，方法和图谱
公开(公告)号：KR1020070112785A
公开(公告)日：2007-11-27
申请号：KR1020077019819
申请日：2006-02-01
申请人： 에이젠코트 바이오사이언스 코오포레이션
发明人： 맥커난,케빈 , 블란차드,알란 , 코틀러,레브 , 코스타,지나
IPC分类号： C12Q1/68 , G06F19/28
CPC分类号： C12Q1/6874 , B82Y15/00 , B82Y30/00 , C12Q1/68 , C12Q1/6837 , C12Q1/6844 , C12Q1/6869 , C12Q2533/107 , C12Q2537/165 , C12Q2565/102 , C12Q2565/137 , C12Q2565/513 , C12Q2565/518 , C12Q2565/537 , G06F19/28
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.
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。在某些实施方案中，所述方法利用含有无碱基残基或受损碱基的延伸探针，并使用适于切割核苷和脱碱基残基之间的连接的试剂和/或适于从核酸中除去受损碱基的试剂。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，测序反应在附着于珠上的模板上进行，所述模板固定在半固体支持物中或半固体支持物上。本发明进一步提供了包含适合用于该方法的硫代磷酸酯键或触发性残基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。本发明还提供了用于制备模板的有效方法，特别是用于并行地进行多个不同模板的排序。本发明还提供了进行连接和切割的方法。本发明还提供了含有配对标签的核酸片段的新文库，以及制备具有多个不同模板（例如，含有配对标签）的微粒和分别对模板进行测序的方法。本发明还提供自动排序系统，流动池，图像处理方法和存储计算机可执行指令（例如，执行图像处理方法）和/或序列信息的计算机可读介质。在某些实施例中，序列信息存储在数据库中。

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