2-甲基异茨醇降解菌阴沟肠杆菌Z5的分离鉴定及BAC文库的构建

Isolation,Identification and BAC Library Construction of an Efficient 2-Methylisoborneol Degradation Bacterium Enterobacter cloacae Z5

从采集到的污水样品中分离得到1株高效降解2-甲基异茨醇(2-methylisoborneol,2-MIB)的菌株Z5,通过16S rDNA序列分析,鉴定为阴沟肠杆菌(Enterobactercloacae)。该菌在以2-MIB为唯一碳源的培养基中能正常生长代谢。经气相色谱-质谱分析,发现该菌对2-MIB的降解率高达89.7%。同时,测定了Z5在不同质量浓度2-MIB中的生长情况,结果表明:当2-MIB质量浓度为16μg/L时,其生长状况与在LB中最为接近。为了克隆降解2-MIB关键酶的编码基因并进一步研究其降解途径,本研究构建了E.cloacaeZ5的全基因组BAC文库,该文库覆盖了大约8倍的基因组,共计810个转化子,外源片段平均大小为40 kb。

A bacterium strain Z5, which can degrade 2-methylisoborneol efficiently, was isolated from sewage samples collected from the sewage treatment plant of Wuhan. This strain was identified as Enterobacter cloacae via the analysis of 16S rDNA. The results of the sole carbon source assay demonstrated that this stain was able to metabolize 2-MIB and the degradation rate was up to 89. 7% determined by GC-MS. The growth of E. cloacae Z5 in 2-MIB at different concentrations were also determined and the growth curve in 16 μg/L 2-MIB was found to he as good as that in LB. To clone the gene coding for the key enzyme catalyzing the degradation of 2-MIB and to further investigate the degradation pathway,the global genome BAC library of E. cloacae Z5 was constructed. There were 810 clones in the library. The average size of inserts and the genome coverage was 40 kb and about 8-folds,respectively.