摘要
以甲基营养细菌Methylobacterium sp.MB200为出发菌株,首先利用质粒转座子(pTnMod-RKm’)构建了目的菌株MB200的部分突变体库,获得能够在双抗性MMII板上生长的突变体共11552株,然后在双抗性MMI板上复筛获得不能够利用甲醇的突变体333株(初步认为是一碳代谢途径被破坏的突变株),为一碳代谢相关基因的克隆研究奠定了基础。随后利用TAIL-PCR技术快速准确地从突变体库中定位了部分与一碳代谢相关的基因,并根据pTnMod-RKm’具有复制起始位点能够快速克隆插入位点侧翼序列的特性克隆到mtdA和mtdB基因的全序列并进行了初步分析。
The partial mutants library of Methylobacterium sp. MB200 was constructed by using plasmid pTnMod-RKm＇ and 11554 mutants were obtained which could survive in MMII containing two antibiotics （Nm and Km）. Rescreening results showed that 333 strains could not use methanol as the sole carbon source,these strains were preliminarily considered that their one carbon metabolism way had been destroyed. This laid the foundation for cloning genes related to one carbon metabolism. The location analysis was carried out by TAIL- PCR, based on the characters of pTnMod-RKm＇ and the full sequence of mtdA and mtdB genes were cloned .
出处
《中国生物工程杂志》
CAS
CSCD
北大核心
2011年第2期50-55,共6页
China Biotechnology
