克雷伯氏菌甘油脱水酶基因在大肠杆菌中的克隆与表达

Cloning and expression of glycerol dehydratase gene from Klebsiella pneumoniae in Escherichia coli

利用PCR技术从克雷伯氏菌 (KlebsiellapneumoniaeATCC4 9790 )总DNA中扩增得到甘油脱水酶 (glyceroldehydratase ,DHAB)基因的DNA片段 ,并将其连接到表达质粒 pSE380 ,携带有重组质粒 pSE dhaB的大肠杆菌JM 1 0 9实现了dhaB基因的表达 ;对含有dhaB工程菌进行表达研究 ,表明工程菌在 37℃ ,以 1 .0mmol LIPTG诱导 5h酶活力即达到 1 1 6 4.1 4U L ,比野生菌酶活力 (1 6 8.6 9U L)提高了 6 .9倍。

Genomic DNAof glycerol dehydratase(DHAB) was used as a template.A segment containing of glycerol dehydratase from c(ATCC49790) was obtianed by PCR amplification and the pSE380 vector combined with the DHAB gene was transformed in JM109. The resulted recombinant Escherichia coli gave a high level expression of DHAB.Some factors were optimized for enzyme overproduction in this experiment.The optimization increased DHAB expression by 6.9 fold in compared to that of K.pneumoniae (168.69U/L) .The enhanced expression of DHAB in recombinant E.coli reached up to 1164.14U/L under the induced condition of 1.0mM IPTG at 37℃for 5 hours.