摘要
以高效产氢细菌哈尔滨产乙醇杆菌的模式菌株YUAN-3作为出发菌株,提取细菌总DNA,经过Sau3AI的不完全酶切,将2.5～5.0kb的片段连接入用BamHI酶切的pUC19质粒,转化入E.ColiDH5α中,构建菌株YUAN-3的基因组文库,得到克隆数接近9×103个,以最相近的梭菌属已报道的基因组平均大小4.6Mb计算,概括了其99%以上的基因组,达到了建库的理论值.随机挑取200个白色菌落进行测序,经分析得到的结果大多数与已经过全序列分析的产氢细菌的假想蛋白相近,其中有49个与其他菌株功能蛋白相似性超过70%的片段,包括叶酰聚谷氨酸合成酶、DNA拓扑异构酶、乙酰转移酶等,同时获得了7个功能蛋白或基因的完整开放阅读框.
The genomic DNA of H2-producing bacterium Ethanoligenens harbinense YUAN-3 was isolated and partially digested by Sau 3A I.The fragments of 2.5-5.0 kb were selected and inserted into BamH I digested plasmid pUC 19.The genomic clone library of strain YUAN-3 was constructed with 9 × 10^3 recombinants,indicating the coverage of clone library tends to be the theoretical value based on a 4.6 Mb average chromosome size of Clostridium,closest to Ethanoligenens harbinense.200 colonies were picked stochastically for sequencing and aligned with the database.The results imply that most of the sequences are similar with the hypothetical protein of hydrogen-producing bacterium whose complete genomes are sequenced,among which 49 sequences have the similarity over 70%,including folylpolyglutamate synthetase,DNA topoisomerase,acetyltransferase and so on,and 7 open reading frames （ORFs） of functional proteins or genes can be obtained.
出处
《哈尔滨工业大学学报》
EI
CAS
CSCD
北大核心
2010年第10期1586-1590,共5页
Journal of Harbin Institute of Technology
基金
国家自然科学基金资助项目(30870037)
城市水资源与水环境国家重点实验室(哈尔滨工业大学)开放课题(HCK201018)
关键词
生物制氢
基因文库
产氢细菌
哈尔滨产乙醇杆菌
biohydrogen； gene library； H2-producing bacterium； Ethanoligenens harbinense；
