长双歧杆菌分泌型表达载体的构建及Tum-5基因的表达

Construction of a Bifidobacterium longum secretory expression vector and expression of gene Tum-5

目的构建能在双歧杆菌内稳定存在并高效分泌表达外源基因的长双歧杆菌质粒表达载体。方法以质粒p BAD/glll为基础,以双歧杆菌内源性阿拉伯糖苷酶的分泌性信号肽取代质粒原有的分泌性信号肽,并将双歧杆菌天然质粒的聚合酶基因克隆入载体中,构建表达载体p BBADs。将人Tum-5基因克隆入载体中,构建质粒p BBADs-Tum-5,电转化长双歧杆菌NCC2705,L-阿拉伯糖诱导表达后,Western Blot鉴定基因表达。结果成功地构建了长双歧杆菌分泌型质粒表达载体,Tum-5基因在双歧杆菌内可以表达。结论构建的表达载体为双歧杆菌用于肿瘤靶向基因治疗奠定了基础。

Objective To construct a stable secretory expression vector of Bifidobacterium longum. Methods Using plasmid pBAD/glll as a basis, replaced the original signal peptide sequence with Bifidobacterium endogenous arabinosidase signal peptide sequence, then the Bifidobacterium plasmid polymerase gene （BPP） was cloned into the vector. The expression vector was named pBBADs. The human Turn-5 gene was cloned into the vector. The plasmid pBBADs-Tum-5 was transformed into B. longum NCC2705 by electroporation. After L-arabinose induc- tion, the expression of Tum-5 in B. longum was detected by Western Blot. Results The secretory expression vector of B. longum was constructed successfully, and Turn-5 gene was expressed in Bifidobacterium. Conclusion The construction of the vector has laid foundation for tumor-targeting gene therapy with B. longum.