Isolation and characterization of a strong promoter element from the Streptomyces ghanaensis phage I19 using the gentamicin resistance gene (aacC1) of Tn1696 as reporter
Isolation and characterization of a strong promoter element from the Streptomyces ghanaensis phage I19 using the gentamicin resistance gene (aacC1) of Tn1696 as reporter
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DOI：
10.1099/00221287-143-5-1503
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发表时间：
1997-05-01
期刊：
影响因子：
--
通讯作者：
Wohlleben, W
中科院分区：
文献类型：
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作者：
Labes, G;Bibb, M;Wohlleben, W
用含有Tn 1696无启动子的氨基糖苷-O-乙酰转移酶I基因（aacC 1）的启动子-探针穿梭质粒（pGL 7011）从加纳链霉菌噬菌体119中分离出在变铅青链霉菌TK 23中具有启动子活性的DNA片段。大肠埃希菌和沙门氏菌对庆大霉素耐药水平分析。aacC 1 mRNA水平在5. Lividans，鉴定了在两个物种中表现出高水平启动子活性的片段（F14）。随后的分析表明，SF 14（F14的亚克隆片段）的启动子活性约为ermEp* 的两倍，ermEp * 是最强的特征性放线菌启动子之一。SF 14含有两个串联的启动子，14-Ip和p14-IIp，分别具有重叠和相邻的-10和-35区。这两种启动子似乎以不同的效率被天蓝色链霉菌A3的主要RNA聚合酶全酶（E sigma（hrdB））识别（2）。
A promoter-probe shuttle plasmid (pGL7011) containing the promoterless aminoglycoside-O-acetyltransferase I gene (aacC1) of Tn1696 was used to isolate DNA fragments from Streptomyces ghanaensis phage 119 that possessed promoter activity in Streptomyces lividans TK23. Analysis of gentamicin (Cm) resistance levels in Escherichia coli and in S. lividans TK23, and of aacC1 mRNA levels in 5. lividans, identified a fragment (F14) that exhibited a high level of promoter activity in both species. Subsequent analysis revealed that the promoter activity of SF14 (a subcloned fragment of F14) was about twice that of ermEp*, one of the strongest characterized adinomycete promoters. SF14 contained two tandemly arranged promoters, 14-Ip and p14-IIp, with overlapping and adjacent -10 and -35 regions, respectively. Both promoters appear to be recognized with different efficiencies by the major RNA polymerase holoenzyme (E sigma(hrdB)) of Streptomyces coelicolor A3(2).