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木薯醇腈酶cDNA的克隆及其表达载体的构建
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【药学论文网】作者：王丹 何浪 王玉明 潘克俭 李维【摘要】 目的 克隆木薯醇腈酶（HNL）cDNA构建表达载体，以实现木薯醇腈酶基因的高效表达。方法 运用 RT-PCR从木薯幼叶组织中扩增出HNL全长cDNA序列，将其克隆至质粒pBluescript SK中进行序列分析，再利用PCR将其再克隆到酵母表达质粒pPIC3.5K上。结果 经测序分析表明，克隆的cDNA片段和文献报道的4个木薯HNL cDNA 相比，核苷酸同源性最高为99.1%，氨基酸同源性最高为98.8%。结论 经双酶切鉴定的结果表明重组表达载体的表达载体构建成功。
【关键词】 木薯；醇腈酶；甲醇酵母；表达载体
Abstract：Objective To study the cloning of the cDNA of α-HNL and the expression vector construction to realize highly efficient expression in NL gene.Methods The long-length sequence of cDNA of α-HNL was amplified by RT-PCR,then it was cloned into pBluescript SK and analylizd in its sequence.Finall,the cDNA was cloned into an expression vector pPIC3.5K by PCR.Results The sequencing result of the cDNA showed that the sequence encoded for the HNL was not fully consistent with those published.The full sequences analysis demonstrated that the highest homology of cDNA sequence is about 99.
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