Alexander Dylan C, Devlin David J, Hewitt Duane D, Horan Ann C, Hosted Thomas J
New Lead Discovery, Schering Plough Research Institute, 2015 Galloping Hill Road, K15-B425-MS4800, Kenilworth, NJ 07033, USA.
Microbiology (Reading). 2003 Sep;149(Pt 9):2443-2453. doi: 10.1099/mic.0.26318-0.
Micromonospora carbonacea var. africana ATCC 39149 contains a temperate bacteriophage, pMLP1, that is present both as a replicative element and integrated into the chromosome. Sequence analysis of a 4.4 kb KpnI fragment revealed pMLP1 att/int functions consisting of an integrase, an excisionase and the phage attachment site (attP). Plasmids pSPRH840 and pSPRH910, containing the pMLP1 att/int region, were introduced into Micromonospora spp. by conjugation from Escherichia coli. Sequence analysis of DNA flanking the integration site confirmed site-specific integration into a tRNAHis gene in the chromosome. The pMLP1 attP element and chromosomal bacterial attachment (attB) site contain a 24 bp region of sequence identity located at the 3' end of the tRNA. Integration of pMLP1-based plasmids in M. carbonacea var. africana caused a loss of the pMLP1 phage. Placement of an additional attB site into the chromosome allowed integration of pSPRH840 into the alternate attB site. Plasmids containing the site-specific att/int functions of pMLP1 can be used to integrate genes into the chromosome.
非洲碳单胞菌变种ATCC 39149含有一种温和噬菌体pMLP1，它既以复制元件的形式存在，也整合到染色体中。对一个4.4 kb KpnI片段的序列分析揭示了pMLP1的att/int功能，包括一种整合酶、一种切除酶和噬菌体附着位点（attP）。含有pMLP1 att/int区域的质粒pSPRH840和pSPRH910通过从大肠杆菌进行接合转移导入到碳单胞菌属中。对整合位点侧翼DNA的序列分析证实了其在染色体中一个tRNAHis基因上的位点特异性整合。pMLP1的attP元件和染色体细菌附着（attB）位点在tRNA的3'端含有一个24 bp的序列相同区域。基于pMLP1的质粒在非洲碳单胞菌变种中的整合导致了pMLP1噬菌体的丢失。在染色体中放置一个额外的attB位点可使pSPRH840整合到另一个attB位点。含有pMLP1位点特异性att/int功能的质粒可用于将基因整合到染色体中。
