蚯蚓纤溶酶在大肠杆菌中的克隆与表达

根据蚯蚓纤溶酶最佳组分的N-末端氨基酸序列设计简并引物,以蚯蚓cDNA为模板,获得该组分蛋白质的部分编码序列(AF432224,GenBank 登陆号).BLAST相似性分析表明,该序列与U25643和U25648的部分序列相似性达91%, 根据5' 和3' 末端保守序列设计引物,经PCR获得全长cDNA编码序列.将该序列插入pTBY-11质粒,转化大肠杆菌,表达蛋白以包含体形式存在.

Cloning and Expression of Earthworm Fibrinolytic Enzyme in E.coli

Degenerate primers were designed according to the partial N-terminal sequence of the Earthworm Fibrinolytic Enzyme (EFE ) component which showing the best fibrinolytic activity. The 540 bp cDNA fragment (AF432224, accession number for GenBank) was amplified by PCR using earthworm cDNA as a template. BLAST similarity analysis showed that the fragment had 91% similarity with U25643 and U25648.Primers were redesigned based on 5' and 3' conservative terminal sequences, and the entire coding sequence of the EFE component was amplified by PCR. The sequence was inserted into the vector pTBY-11 and transformed into E.coli. The protein was expressed as inclusion bodies.