日本血吸虫溶血磷脂酶基因真核表达载体的构建、表达与鉴定

Gong-si Fang, Yong Yao, Li-wen Wang, Xue-long Wang
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引用次数: 0

摘要

目的从日本血吸虫成虫cDNA中扩增日本血吸虫溶血磷脂酶基因(Sjl539),并将其亚克隆到真核表达载体pcDNA3.1(+)中表达重组抗原并进行免疫原性分析。方法采用RT-PCR法提取日本参总RNA,生成cDNA。扩增Sj1539基因。在pGEM-T中插入扩增后,将该DNA片段亚克隆到真核表达载体pcDNA3.1(+)中。将重组质粒转染人宫颈癌细胞株(Hela细胞),用Westernblotting法鉴定表达产物。结果PCR产物大小约为684 bp。经双酶切重组质粒和PCR证实,sj1539基因已成功插入。结果表明,表达产物能与s反应。经Western blotting检测日本血吸虫感染兔血清,分子量约为25×103。结论建立了Sj1539基因真核表达载体,获得了表达产物。
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Construction,expression and identification of eukaryotic expression vector carrying Schistosoma japonicum gene coding lysophospholipase
:Objective Schistasomajaponicum(S.japonicum)lysophospholipase gene(Sjl539)from cDNA of S japonicum adult wormswas amplified and subcloned into eukaryotic expression vector pcDNA3.1(+)for expression ofrecombinant antigen and immunogenicity analysis.Methods Total RNA of S.japonicum wasextracted to generato cDNA by RT-PCR.The Sj1539 gent was amplified.The DNA fragment wassubcloned into eukaryofic expression vector pcDNA3.1(+)following insertion andamplification in pGEM-T.The recombinant plasmid was transfected into human cervicalcarcinoma cell strain(Hela cells)and expression products were identified by Westernblotting.Results The size of PCR product was approximately 684 bp.It was confirmed thatSj1539 gene had been inserted successfully by the recombinant plasmid digested with twoenzymes and PCR.It was verified that the expression product could react withS.japonicum-infected rabbit serum by Western blotting and the molecular weight wasapproximately 25×103.Conclusions The eukaryotie expression vector carrying Sj1539 genehas been established and the expression product has been obtained.
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来源期刊
中华地方病学杂志
中华地方病学杂志 我国对人类健康危害特别严重的地方性疾病:克山病、大骨节病、碘缺乏病、地方性氟中毒、地方性砷中毒、鼠疫、布鲁氏菌病、寄生虫、新冠肺炎等疾病,同时还报道多发性自然疫源性疾病。
CiteScore
1.60
自引率
0.00%
发文量
8714
期刊介绍: The Chinese Journal of Endemiology covers predominantly endemic diseases threatening health of the people in the areas affected by the diseases including Keshan disease, Kaschin-Beck Disease, iodine deficiency disorders, endemic fluorosis, endemic arsenism, plague, epidemic hemorrhagic fever, brucellosis, parasite diseases and the diseases related to local natural and socioeconomic conditions; and reports researches in the basic science, etiology, epidemiology, clinical practice, control as well as multidisciplinary studies on the diseases.
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