弗氏柠檬酸细菌酪氨酸酚解酶基因在大肠杆菌中的克隆与表达

以基因组DNA为模板，利用PCR技术从弗氏柠檬酸细菌（Citrobacter freundii)中扩增得到含有酪氨酸酚解酶基因的DNA片段，定向连续到质粒pUC118上，得到重组质粒pTPL，将此重组质粒转化到受体菌E.colXL-1-Blue MRF′中，通过蓝白斑鉴定挑出阳性菌株。从此阳性菌株中提取质粒pTPL并将此质粒转入到E.coliJM109中，用E.coliJM109（pTPL)制备高活性的酪氨酸酚解酶。对质粒稳定性的研究表明，E.coliJM109（pTPL)在无选择压力下37℃连续培养50代以上，质粒丢失率仅有15％，说明质粒基本稳定。

Cloning and expression of tyrosine phenol lyase

Genomic DNA of tyrosine phenol lyase was used as a template. A segment containing gene of tyrosine phenol lyase from Citrobacter freundii was gained by PCR amplification and cloned into pUC118 to obtain recombinant plasmids pTPL, then the plasmid was transformed into E.coli XL 1 Blue MRF'. The positive strains were obtained by the determination of blue white plot. The plasmid pTPL was extracted from the positive strains and transformed into E. coli JM109. The cells containing high tyrosine phenol lyase activity were prepared with E.coli JM109(pTPL). The study of plasmid stability indicated the loss rate of plasmid was only 15 percent after cultured 50 generations in the medium without ampicillin, which showed that the plasmid was basically stable.