发明授权
- 专利标题: Techniques for producing site-directed mutagenesis of cloned DNA
- 专利标题(中): 用于产生克隆DNA定点诱变的技术
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申请号: US764085申请日: 1991-09-23
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公开(公告)号: US5284760A公开(公告)日: 1994-02-08
- 发明人: Stephen M. Feinstone , Czeslaw Wychowski , Jonathan E. Silver , Suzanne U. Emerson
- 申请人: Stephen M. Feinstone , Czeslaw Wychowski , Jonathan E. Silver , Suzanne U. Emerson
- 专利权人: Feinstone Stephen M,Czeslaw Wychowski,Silver Jonathan E,Emerson Suzanne U
- 当前专利权人: Feinstone Stephen M,Czeslaw Wychowski,Silver Jonathan E,Emerson Suzanne U
- 主分类号: C07K14/10
- IPC分类号: C07K14/10 ; C07K14/105 ; C12Q1/68 ; C12N15/10
摘要:
A method is described whereby new cDNA or RNA sequences can be introduced into or substituted for cDNA in any chosen position without specific sequence requirements using the polymerase chain reaction (PCR). The method entails the use of primers which are complementary to the 3' and 5' ends of the desired sequence to be inserted as well as to the 3' and 5' ends of the chosen site of insertion in the acceptor molecule. The desired sequence is amplified by PCR such that single stranded fragments are produced. The single stranded fragment of the desired sequence is then annealed to a single stranded acceptor molecule at the site of insertion and extended to produce a double stranded molecule. The double stranded molecule is then separated into two strands which are identical except that one of the strands contains the desired sequence inserted at the chosen site. A second double stranded molecule is then generated.
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