发明名称 |
Method for cloning and producing the SnaBI restriction endonuclease and purification of the recombinant SnaBI restriction endonuclease |
摘要 |
The methylase selection method was used to clone the SnaBI methylase gene (snaBIM) from Sphaerotilus natans (ATCC 15291). An active SnaBI methylase was cloned in E. coli using pSnaBI-2, a pUC19 derivative containing two SnaBI sites. Because methylase and restriction genes are usually located alongside each other in a restriction-modification systems, efforts were made to clone the downstream DNA by inverse PCR. Inverse PCR cloned the missing portion of the SnaBI endonuclease and also identified a control, or C, protein. The two open reading frames snaBIR (ORF1) and snaBIC (ORF2) converged toward the SnaBI methylase gene (ORF). Attempts to establish a snaBIR-recombinant plasmid expressing the SnaBI endonuclease in E.coli modified with SnaBI methylase failed. Overexpression of the SnaBI endonuclease in E. coli required the use of the heterospecific BsaAI methylase.
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申请公布号 |
US6025179(A) |
申请公布日期 |
2000.02.15 |
申请号 |
US19980143776 |
申请日期 |
1998.08.31 |
申请人 |
NEW ENGLAND BIOLABS, INC. |
发明人 |
LUNNEN, KEITH D.;KONG, HUIMIN;WILSON, GEOFFREY G |
分类号 |
C12N15/09;C12N1/15;C12N1/19;C12N1/21;C12N5/10;C12N9/10;C12N9/16;C12N9/22;C12N15/52;C12N15/55;(IPC1-7):C12N9/22 |
主分类号 |
C12N15/09 |
代理机构 |
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