发明名称 |
Methods of using improved polymerases |
摘要 |
This invention provides for methods of sequencing and performing polymerase reactions using an improved generation of nucleic acid polymerases. The improvement is the fusion of a sequence-non-specific nucleic-acid-binding domain to the enzyme in a manner that enhances the processivity of the polymerase. |
申请公布号 |
US9139873(B2) |
申请公布日期 |
2015.09.22 |
申请号 |
US201313924353 |
申请日期 |
2013.06.21 |
申请人 |
Bio-Rad Laboratories, Inc. |
发明人 |
Wang Yan;Vander Horn Peter;Xi Lei |
分类号 |
C12N9/12;C12Q1/68;C07K14/195;C12N15/10;C12N15/62;C12P19/34 |
主分类号 |
C12N9/12 |
代理机构 |
Kilpatrick Townsend & Stockton LLP |
代理人 |
Kilpatrick Townsend & Stockton LLP |
主权项 |
1. A method of sequencing a target nucleic acid in an aqueous solution using an improved DNA polymerase, the method comprising:
(a) contacting the target nucleic acid with a thermally stable DNA polymerase, wherein the DNA polymerase is joined to a sequence non-specific double-stranded nucleic acid binding domain that comprises at least 75% amino acid sequence identity to the Sso7d amino acid sequence set forth in SEQ ID NO:2, and enhances the processivity of the DNA polymerase compared to an identical DNA polymerase not having the sequence non-specific double-stranded nucleic acid binding domain fused to it, and
wherein the solution is of a composition that permits the sequence non-specific double-stranded nucleic acid binding domain to bind to the target nucleic acid and the polymerase domain to extend a primer that is hybridized to the target nucleic acid sequence; (b) incubating the solution under conditions in which the primer is extended by the DNA polymerase; and (c) identifying the nucleotides that are incorporated into the synthesized nucleic acid strand upon the template-dependent extension of the primer by the DNA polymerase, thereby sequencing the nucleic acid. |
地址 |
Hercules CA US |