摘要
用启动子探针质粒ｐＩＪ４８６从麦迪霉素产生菌（Ｓ．ｍｙｃａｒｏｆａｃｉｅｎｓ）中克隆到１个具启动功能的ＨｉｎｄＩＩＩ－ＨｉｎｄＩＩＩ２．０ｋｂ片段，含该片段的重组质粒ｐ４Ｈ２转化子在ＭＭ基本培养基上对卡那霉素（Ｋｍ）抗性可达５００μｇ／ｍｌ以上。亚克隆缺失分析结果表明，该片段不同部分的缺失对启动活性有不同程度的影响，说明它具有较复杂的转录调控机制。ＤＮＡ序列分析结果显示，该片段含有１９８４个核苷酸，其Ｇ＋Ｃ％为４７．７％，不存在典型的链霉菌的可读框；进一步的分析发现，其６５０ｂｐ、１１５０ｂｐ和１５００ｂｐ区域分别与麦迪霉素酮基还原酶基因等链霉菌基因的启动子序列具有同源性；在５２０－５７０ｂｐ区域与大肠杆菌ｔＲＮＡ基因上游激活序列（ＵＡＳ）有较好的同源性，提示链霉菌中可能存在可增强基因转录的ＤＮＡ元件。
A HindIII-HindIII 2.0 kb DNA fragment containing promoter activity has been cloned from medicamycin producing strain(S. mycarofaciens 1748),using promoter-probe plasmid vector pIJ486. Transformants of recombinant plasmid p4H2 containing this fragment were resistant to Km in the level above 500μg/ml in MM medium. The result of subcloning deletion analysis indicated that different part of deletion of this fragment would show various extent of effect on the promoter activity. A rather complicated transcriptional regulatory mechanism was suggested. DNA sequence analysis revealed that this fragment consisted of 1984 nucleotides with 47.7% of G+C content. No typical ORF of Streptomyces gene was found. Certain regions showed some homologies with promoters of Streptomyces genes. In 520-570bp region a sequence similiar to the upstream activator sequence (UAS) of E. coli tRNA was discovered, This implied that a possible 'enhancer' element was involved in Streptomyces gene transcription.
基金
国家自然科学基金
关键词
链霉菌
启动子
DNA序列
结构
功能
Streptomyces promoter, DNA sequence, Upstream activator sequence
