安徽农业科学2017，Vol.45Issue(36)：131-134,153,5.
一株石油烃降解菌TY22烷烃羟化酶基因的克隆及功能研究
Cloning and Functional Analysis of Alkane Hydroxylase Gene of a Petroleum Hydrocarbon Degrading Bacteria TY22
摘要
Abstract
[Objective] To clone and sequence the alkane hydroxylase gene alkB of a petroleum hydrocarbon degrading bacteria TY22,then to clone alkB into express vector for expressing and identifying its function.[Method] First,use PCR amplification to get partial sequence of 1236 bp alkB gene,then obtain the complete CDS sequence by chromosome walking techniques.Second,clone alkB gene into pET21b( +) vector,and transform the recombinant plasmid into E.coli BL21(DE3) for expressing.Third,clone alkB gene into pCom8 vector,and transform the recombinant plasmid into E.coli GEc137(pGEc47△B) and Pseudomonas fluorescens KOB2Δ1 respectively for gene function complementation test.[Result] We got the complete CDS sequence of 1236 bp alkane hydroxylase gene through chromosome walking techniques ( GenBank Accession:KU867870).The expression strain had the most expression with 0.1 mmol/L IPTG,30 ℃culture for 6 hours,relative molecular mass 46 kDa was also in line with expectation.Gene function complementation test showed that the recombinant strains can grow in the medium with n-alkanes C12～C16 as sole carbon source.[Conclusion] Alkane hydroxylase gene alkB complete CDS sequence length of strain TY22 was 1236 bp.It can oxidize n-alkanes with chain length C12 ～C16 ,and efficient expression in E.coli BL21(DE3).关键词
石油烃降解菌/烷烃羟化酶/表达/功能Key words
Petroleum hydrocarbon degrading bacteria/Alkane hydroxylase/Expression/Function分类
农业科技引用本文复制引用
唐赟,敬珊,邹书珍,冀黎骏,杨雪,余小丽..一株石油烃降解菌TY22烷烃羟化酶基因的克隆及功能研究[J].安徽农业科学,2017,45(36):131-134,153,5.基金项目
四川省科技厅应用基础研究项目(2014JY0196) （2014JY0196）
四川省南充市科技局重点项目(16YFZJ0040). （16YFZJ0040）
