链霉菌质粒pIJ101最小复制子的鉴定

Identification of the minimal replicon of the streptomycete plasmid pIJ101.

作者信息

Zaman S, Richards H, Ward J

机构信息

Department of Biology, University College London, United Kingdom.

出版信息

Plasmid. 1993 Jan;29(1):57-62. doi: 10.1006/plas.1993.1007.

DOI:10.1006/plas.1993.1007

PMID:8441769

Abstract

The minimal replicon of pIJ101 was defined to a 2.0-kb BalI-SacII fragment containing the rep ORF and a non-coding region of DNA, but not orf56 (pQR435). This non-coding region of DNA was found to be crucial for plasmid replication in Streptomyces lividans and may contain either the origin of replication or the rep promoter (or both). A 1.4-kb NotI-SacII fragment containing only the rep ORF produced a 54-kDa protein in vitro when expressed from the lacZ promoter. pQR435 was structurally unstable in S. lividans and accumulated single-stranded (ss)DNA. The insertion of the site for second-strand initiation (ssi) into pQR435 did not prevent the plasmid from rearranging or from accumulating ssDNA.

摘要

pIJ101的最小复制子被定义为一个2.0 kb的BaI - SacII片段，其包含rep开放阅读框（ORF）和一段DNA非编码区，但不包含orf56（pQR435）。发现该DNA非编码区对于变铅青链霉菌中的质粒复制至关重要，并且可能包含复制起点或rep启动子（或两者）。当从lacZ启动子表达时，一个仅包含rep ORF的1.4 kb NotI - SacII片段在体外产生了一种54 kDa的蛋白质。pQR435在变铅青链霉菌中结构不稳定，并积累单链（ss）DNA。将第二链起始位点（ssi）插入pQR435并不能阻止质粒重排或积累ssDNA。