大肠杆菌色氨酸酶基因的克隆与表达

Molecular Cloning and Expression of Tryptophanase Gene of Escherichia coli

摘要: 应用ＰＣＲ技术从Ｅ．ｃｏｌｉＪＭ１０５中扩增出长约１．４Ｋｂ的色氨酸酶基因，将其插入高表达载体ｐＥＴ３ａ的ＮｄｅＩ／ＢａｍＨＩ位点，转化Ｅ．ｃｏｌｉＢＬ２１（ＤＥ３），构建高产色氨酸酶基因工程菌。ＳＤＳ－ＰＡＧＥ电泳和薄层扫描表明，工程菌色氨酸酶的表达量占细胞总可溶性蛋白的６９．８％。酶活测定结果表明，７株工程菌色氨酸酶的活力比宿主菌均有不同程度的提高，其中ＷＷ－１１号比宿主菌高１６倍。

Abstract: kb DNA fragment was obtained from E. coli JM 105 by PCR. This fragment was inserted into NdeI/BamHI sites of pET3a. The recombinant plasmids were transformed into host strain E. coli BL 21(DE3). High level expression of tryptophanase genetic engi