依尼奥小单孢菌抗性基因sisR的克隆研究

从产西索米星的依尼奥小单孢菌中克隆高抗性新基因sisR，借助计算机设计PCR引物，从西索米星的产生菌依尼奥小单孢菌的染色体DNA中，经PCR扩增获得DNA序列长度不等的DNA片段。将这些DNA片段克隆至pUC19载体质粒并导入大肠杆菌，从中筛选到5个抗西索米星的转化子(其中一个命名为sisR)显示对西索米星的高抗性(超过1000μg/mL)。经DNA测序并通过互联网Biast比对，确认对该抗性负责的DNA片段是一个未见报道的新基因。

Study on Cloning of Sisomicin-resistant Gene ( sisR ) from Micromonospora inyoensis

A new sisomicin resistance gene sisR was cloned from sisomicin-producing Micromonospora inyoensis. The sisR fragment was obtained by PCR amplification. The primer pairs were designed based on grm gene sequence from gentamicin-producing Micromonospora purpurea. The template DNA was isolated from Micromonospora inyoensis. A series of different DNA fragments were amplified by PCR, which were sub-cloned to vector pUC19 for further identification. It was found that five specific transformants containing target DNA fragments could resist high concentrations of sisomicin (over 1000 microg/mL sisomicin). One of them designated as sisR, was then sequenced and the alignment among sisR and other related genes showed that sisR gene differs from any known genes. It was concluded that sisR gene is a sequence that has not been reported so far.