地中海拟无枝菌酸菌U32染色体特性的脉冲场电泳分析

地中海拟无枝菌酸菌(Amycolatopsis mediterranei)U32是产力复霉素SV的工业生产菌株。采用脉冲场电泳分析发现，地中海拟无枝菌酸菌U32仅有一条约10 Mb的线性染色体, 没有内源性质粒。利用Southern杂交法，对11个编码力复霉素生物合成、相关初级、次级代谢关键酶以及调控蛋白的基因，在U32染色体DNA的PshBI酶切片段上进行了定位。分析发现在一条长度约700kb的PshBI酶切片段上，分别存在着力复霉素合成基因簇(rif)、氮代谢的亚硝酸还原酶小亚基基因(nasD)、衔接初级与次级代谢的甲基丙二酰变位酶基因(mcm)、脂肪酸代谢的乙酰辅酶A羧化酶生物素载体蛋白基因(accA)以及一套核糖体RNA转录单元。同时还发现U32至少有5套核糖体RNA转录单元。其余定位的基因均只出现单一杂交信号。

Characterization of Amycolatopsis meditteranei U32 Chromosome by Pulse-field Gel Electrophoresis

Amycolatopsis meditteranei U32 is an industrial strain with high yield of rifamycin SV. Its chromosome structure was characterized by pulse-field gel electrophoresis. The topological structure of U32 chromosome was linear and its genome size was about 10 Mb. There was no internal plasmid found. Gene encoding several enzymes involved in rifamycin bio-synthesis and relevant primary metabolism as well as protein related to metabolic regulation were localized by Southern blot. The results revealed that there were at least five copies of rrn operon in U32. The rifamycin bio-synthesis gene cluster was located on one about 700 kb Psh BI fragement where several other important genes related to primary and secondary metabolism were also situated. They were the nasD gene, the mcm gene encoding methylmalonyl CoA mutase, the acc A gene encoding acethyl CoA carboxylase biotin carrier protein and one copy of the rrn operons.