两种血吸虫病疫苗候选分子在原核细胞的融合表达

THE FUSION EXPRESSION OF TWO CANDIDATE OF SCHISTOSOMA JAPONICUM VACCINE IN PROKARYOTIC CELL

目的 在原核细胞中表达 Sj GST和 Sj14- 3- 3的融合蛋白。 方法 RT- PCR法从日本血吸虫成虫总 RNA中扩增出 Sj14- 3- 3基因 ,亚克隆至原核表达载体 p GEX- 4T- 1,并转化 E.coli BL2 1 ,IPTG诱导表达 ,Western- blot鉴定表达产物。 结果 扩增产物约为 76 5 bp,重组质粒经 Bam H 和 Xho 双酶切后可得到一与 PCR产物大小相同的 DNA片段 ,经 IPTG诱导后在 5 5 ku附近出现一新增蛋白条带 ,Western- blot结果显示 ,表达产物可被兔抗鼠 14- 3- 3ε多克隆抗体识别。 结论 在原核细胞融合表达 Sj GST和 Sj14- 3- 3成功。

Objective To express the fusion protein SjGST and Sj14-3-3 in prokaryotic cell. Methods The Sj14-3-3 gene was amplified from the total RNA of Schistosoma japonicum by RT-PCR, and subcloned into the prokaryotic expression vector pGEX-4T-1. The recombinant plasmid pGEX-4T-1-Sj14-3-3 was transformed into E. coli BL 21, and induced by IPTG. By western-blot, the fusion protein was identified. Results The PCR product was about 765 bp. A DNA fragment same size as the PCR product appears after the recombinant plasmid was digested with BamHⅠand XhoⅠ, A new protein band of 55 ku emerges after the positive clone was induced by IPTG. Result of western-blot indicated that the fusion protein can be recognized by the polyclone antibody against 14-3-3ε. Conclusion The expression of the fusion protein of SjGST and Sj14-3-3 succeded in prokaryotic cells.