摘要
包涵体的形成与外源基因在大肠杆菌中的表达量高度相关 ,在适当的范围内 ,降低hbFGF在表达宿主BL2 1 (DE3)plysS中的表达 ,成为实现高可溶性表达的关键。在不改变氨基酸序列的条件下 ,对hbFGF高表达菌株突变重组子起始密码ATG下游前 3个密码子的摇摆碱基进行随机回复突变 ,共有 7种组合 ,合成引物PCR扩增后 ,克隆至表达载体pET 3c ,将重组子转导BL2 1 (DE3)plysS后 ,IPTG诱导表达 ,发现其中 1株有较高可溶性和活性的菌株。可见部分降低外源蛋白的表达量可以避免与减少包涵体的形成。
The formation of inclusion body is correlated with the overexpression of foreign proteins in Escherichia coli . Controlling of the expression level seems to be critical to increase the soluble component. With the prerequisite of no alterations in amino acid sequence, the wobble bases of the first 3 codons downstream the initiation codon ATG of a hbFGF high-expression mutation, established previously, were changed into the bases of the native hbFGF sequences. Seven primers were synthesized for retro-mutations, after PCR were undertaken, the products were cloned into pET-3c, and recombinants were transformed into BL21(DE3)plysS for expression. One strain with high solubility and bioactivity were identified, indicating that restriction of the expression level of recombinant protein is helpful for high-solubility.
出处
《微生物学报》
CAS
CSCD
北大核心
2003年第4期448-452,共5页
Acta Microbiologica Sinica
基金
国家"8 63计划"资助项目 (Z1 8 0 3 2 9)~~
