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目的 构建日本血吸虫中国大陆株SjF1原核表达重组质粒,并进行免疫保护效果测定。方法 用聚合酶链反应(PCR)法将SjF1基因从已剪切阳性克隆中扩增出来,克隆入原核表达载体pTWIN1上intein2的N端,经PCR和限制性酶切筛选阳性重组子。将阳性重组子转化大肠杆菌,在低温和低IPTG浓度下诱导表达可溶性重组融合蛋白。经鉴定后分别以FCA作佐剂经皮下免疫和以壳聚糖作佐剂经黏膜免疫昆明鼠,观察诱导产生的减虫和减卵效果。感染前采血和唾液用ELISA法检测抗体。结果 成功克隆并表达了rSjF1。rSjF1以FCA作佐剂经皮下免疫获得了29.92％的减虫率和51.08％的减卵率;rSjF1以壳聚糖作佐剂经滴鼻免疫获得了25.74％的减虫率和44.04％的减卵率。结论 获得了rSjF1,该蛋白以FCA作佐剂经皮下免疫和以壳聚糖作佐剂经黏膜免疫均能诱导小鼠产生部分抗血吸虫感染的保护力。
Objective To construct the recombinant plasmid of SjF1 from Chinese mainland Schistosoma japonicum and evaluate the effect of immunoprotection. Methods The SjF1 gene was amplified from the cloned positive clones by polymerase chain reaction (PCR) and cloned into the N terminus of intein2 on the prokaryotic expression vector pTWIN1. The positive recombinant was screened by restriction endonuclease and restriction enzyme digestion. Positive recombinants were transformed into E. coli to induce the expression of soluble recombinant fusion proteins at low and low IPTG concentrations. After being identified, the mice were immunized subcutaneously with FCA as an adjuvant and mucosal immunized Kunming mice with chitosan as an adjuvant to observe the effects of de-worming and reducing ovulation. Pre-infection blood and saliva were detected by ELISA antibody. Results The successful cloning and expression of rSjF1. rSjF1 achieved an ECM of 29.92% and an egg reduction rate of 51.08% with adjuvant FCA as an adjuvant. An immunosuppressive rate of 25.74% and a reduction of 44.04% for rSjF1 were obtained by intranasal immunization with chitosan as an adjuvant Egg rate. CONCLUSIONS: rSjF1 was obtained. The protective effect of rSjF1 on partial anti-schistosome infection induced by mucocutaneous immunization with FCA as an adjuvant and mucosal immunization with chitosan as adjuvant was obtained.