日本血吸虫FKBP12基因的原核克隆表达及表达产物的纯化

CLONE AND IMMUNOLOCALIZATION OF SCHISTOSOMA JAPONICUM FKBP12 GENE

目的 扩增、原核克隆和表达日本血吸虫 (Sj) FKBP12基因全长 c DNA,并进行表达产物的纯化。方法 从成虫 RNA中 RT- PCR扩增 Sj FKBP12基因完整的编码阅读框后 ,将其克隆入p GEX- 4 T- 1原核表达载体中 ,IPTG诱导高表达后用亲和层析柱对表达产物进行纯化。结果 将 SjFKBP12基因成功克隆入 p GEX- 4 T- 1表达载体中 ;诱导表达并成功纯化出重组 FKBP12蛋白。结论 成功克隆了 Sj FKBP12基因 ,并表达和纯化得到了 FKBP12蛋白 ,为研究 FKBP12的功能及其进行疫苗等研究奠定了基础。

Objective To clone and express Schistosoma japonicum (Sj) FKBP12 gene, and then purify the expressed protein. Methods FKBP12's complete coding sequence(CDS) from adult worm mRNA was amplified by RT-PCR. cDNA of FKBP12 gene was cloned into the pGEX-4T-1 vector. The recombinant plasmid was transformed into [WT5”BX]E.coli BL21(DE3) and expressed, then the expressed products in E.coliwere purified by the affinity chromatography column.Results FKBP12 gene cDNA sequence was obtained and then successfully cloned into pGEX-4T-1 vector. FKBP12 fusion protein was expressed after it was induced by IPTG and purified with 26 kDa GST at the N terminus. Conclusion Sj FKBP12 gene is successfully cloned, expressed and the fusion protein purified, which give the basis for further study especially as a vaccine.