目的构建弓形虫致密颗粒蛋白7(GRA7)基因重组质粒并在大肠埃希菌中进行表达。方法根据GRA7基因序列设计合成引物,用PCR方法从弓形虫基因组DNA中扩增GRA7基因片段,再克隆到pGEX-4T载体中,重组质粒经酶切、PCR鉴定并测序;重组质粒在大肠埃希菌BL21(DE3)中诱导表达,表达产物经SDS-PAGE分析并纯化,Western blot分析其反应原性。结果 GRA7基因PCR产物大小约为714bp,与预期相符;重组质粒经酶切及PCR鉴定构建成功,测序结果与已知序列吻合;重组质粒转化菌经IPTG诱导后表达的GRA7融合蛋白分子质量单位约为53ku,该蛋白可被His标签抗体识别。结论成功重组了弓形虫GRA7基因,表达蛋白具有反应原性,为弓形虫诊断抗原试剂盒的制备奠定了基础。 Objective To construct the recombinant plasmid of Toxoplasma gondii GRP7 gene and express it in Escherichia coli. Methods According to the sequence of GRA7 gene, a synthetic primer was designed. The GRA7 gene fragment was amplified from the genomic DNA of Toxoplasma gondii by PCR and cloned into pGEX-4T vector. The recombinant plasmid was identified by PCR and identified by PCR. The recombinant plasmid was expressed in Escherichia coli The expression in E. coli BL21 (DE3) was induced. The expressed product was analyzed by SDS-PAGE and purified. The reaction product was analyzed by Western blot. Results The size of PCR product of GRA7 gene was about 714bp, which was consistent with the expectation. The recombinant plasmid was successfully constructed by restriction enzyme digestion and PCR. The sequencing result was consistent with the known sequence. The molecular mass unit of GRA7 fusion protein expressed by IPTG 53ku, this protein can be His-tag antibody recognition. Conclusion The GRA7 gene of Toxoplasma gondii was successfully recombined, and the expressed protein was reactive. It laid the foundation for the preparation of Toxoplasma gondii antigen kit.