大肠杆菌-双歧杆菌穿梭表达载体的构建及内皮抑素基因的表达

Constraction of a Escherichia coli-Bifidobacterium longum Shuttle Expression Vector and Expression of Human Endostatin Gene

目的:构建大肠杆菌-长双歧杆菌穿梭表达载体,并通过此载体使人内皮抑素基因在大肠杆菌和长双歧杆菌中得到表达。方法:以质粒pDG7、pBCSK(+)、pET-9C为基础,构建大肠杆菌-长双歧杆菌穿梭表达载体pET-1128,并将人内皮抑素基因插入到新构建的表达载体中,分别转化大肠杆菌BL21(DE3)和长双歧杆菌NQ-1501。诱导表达,表达产物经SDS-PAGE和WesternBlot鉴定。结果:成功构建了大肠杆菌-双歧杆菌穿梭载体,人内皮抑素基因在大肠杆菌和长双歧杆菌中均可表达。结论:构建的穿梭载体为今后用双歧杆菌作为生理菌载体进行肿瘤的基因治疗奠定了基础。

Objective： To construct Escherichia coli-Bifidobacterium longum shuttle expression vector. Methods： The E. coli-B.longum shuttle expression vector pET-1128 was constructed based on plasmids of pDG7, pBCSK（＋） and pET-9C. Then the plasmid pET-1128 with human endostatin gene was used to transform B.longum NQ-1501 and E.coli BL21 （DE3）. The recombinant strains were induced separately. SDS-PAGE and Western Blot were used to verify the product of expression of endostatin gene. Results： The expression products of human endostatin gene was both observed in E.coli and B.longum. Conclusions： The E.coli-B.longum shuttle vector was constructed successfully. This study laid the base of using Bifidobacterium as cancer gene therapy vector.