摘要
采用寡核苷酸介导的定向点突变法，对天蚕抗菌肽Ｃｅｃｒｏｐｉｎ Ｂ基因１３ 位甲硫氨酸（ Ｍｅｔ） 诱变为亮氨酸（Ｌｅｕ） 或缬氨酸（Ｖａｌ） ，保留１ 位上的Ｍｅｔ，诱变后的Ｃｅｃｒｏｐｉｎ ＢＤＮＡ序列分析证明产生了预期的点突变。将Ｃｅｃｒｏｐｉｎ Ｂ 突变体基因与ｐＧＥＸ４Ｔ２ 融合表达载体中的谷胱甘肽转移酶（ＧＳＴ） 基因融合，在Ｅ．ｃｏｌｉ 中表达，当ＩＰＴＧ 诱导３０ｍｉｎ 后，工程菌的数量开始减少，然后逐渐恢复正常。说明Ｃｅｃｒｏｐｉｎ Ｂ 突变体与ＧＳＴ 基因融合表达后仍然具有很强杀伤原核细胞的作用。
Using the method of oligonucleotide mediated site directed mutagenesis, We have mutated the Met13 of Cecropin B gene for Leu13 or Val13. The DNA sequencing analysis of CecropinB mutant displayed a expecting site directed Mutagenesis. The fusion expression vetor pGEX 4T 2 contains GST gene, which encodes Glutathione S Transferese fused to modified Cecropin B gene and expressed in E.coli TG1. After 30min of IPTG induction, a large number of engineering bacteria were dead. The results demonstrated the fusion protein of GST CecropinB mutant have still killed the protocaryon cell.
出处
《药物生物技术》
CAS
CSCD
1999年第4期193-197,共5页
Pharmaceutical Biotechnology
基金
国家863 高技术基金!项目(101010203)
