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中国生物工程杂志

CHINA BIOTECHNOLOGY
中国生物工程杂志
中国生物工程杂志  2010, Vol. 30 Issue (05): 36-42    
研究报告     
日本血吸虫三价DNA疫苗pVIVO2SjFABP/Sj26.SjGAPDH的构建及其免疫保护作用评价
朱路1,2,龙全科1,2,刘海峰1,2,鲁明波1,2,胡媛3,石佑恩3,余龙江1,2**
1.分子生物物理教育部重点实验室 武汉 430074
2.华中科技大学生命科学与技术学院 武汉 430074
3.华中科技大学同济医学院 武汉 430030
Construction and Immuno-protection Evaluation of Trivalent-epitope DNA Vaccine pVIVO2-SjFABP/Sj26.SjGAPDH
ZHU Lu1,2,LONG Quan-ke1,2,LIU Hai-feng1,2,LU Ming-bo1,2 ,HU Yuan3,SHI You-en3,YU Long-jiang1,2
1.Key Laboratory of Molecular Biophysics, Ministry of Education, Wuhan 430074, China
2.College of Life Science and Technology, Huazhong University of Science and Technology, Wuhan 430074, China
3.Tongji Medical College, Huazhong University of Science and Technology, Wuhan 430030, China
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摘要：

目的:构建含有日本血吸虫中国大陆株脂肪酸结合蛋白(SjFABP)，3-磷酸甘油醛脱氢酶(SjGAPDH)，26kDa谷胱甘肽S转移酶(Sj26)的三价DNA疫苗，并通过药理实验评价其抗血吸虫感染的免疫保护作用。 方法:  以血吸虫成虫RNA为模板制备cDNA第一链，RT-PCR扩增得到抗原基因片段，再通过重组PCR技术，将Sj26和SjGAPDH融合为Sj26.SjGAPDH基因。将SjFABP和Sj26.SjGAPDH分别克隆入pVIVO2的mcs1和mcs2，获得重组质粒pVIVO2-SjFABP/Sj26.SjGAPDH。瞬时转染MCF-7细胞，通过逆转录PCR（RT-PCR）检测抗原基因在mRNA水平的表达，通过间接荧光免疫（IIF）检测抗原基因在小鼠体内抗原水平表达，验证了DNA疫苗的有效性；并免疫小鼠后进行免疫保护性初步试验。结果:  经过酶切、测序及体内外表达验证，该三价DNA疫苗pVIVO2-SjFABP/Sj26.SjGAPDH构建成功；该三价DNA疫苗在小鼠抗血吸虫感染中诱发减虫率和减卵率达到58.6%和59.8%。结论:  该三价DNA疫苗组具有比单价和双价DNA疫苗组更加良好的免疫保护作用，为日本血吸虫DNA疫苗的研制奠定了基础。
关键词： 日本血吸虫;  脂肪酸结合蛋白;  3-磷酸甘油醛脱氢酶;  26kDa谷胱甘肽S转移酶;  三价DNA疫苗    
Abstract:

Objective: To construct trivalent-epitope DNA vaccine encoding SjFABP, SjGAPDH and Sj26, and evaluate the protective immunity efficacy against Schistosoma japanicum by pharmacological trials. Methods: The monovalent antigen gene SjFABP, SjGAPDH and Sj26 were obtained by RT-PCR. Sj26 and SjGAPDH are spliced into covalent fusion gene of Sj26.SjGAPDH by recombinant PCR. pVIVO2-SjFABP/Sj26.SjGAPDH is constructed by subcloning the SjFABP and Sj26.SjGAPDH into mcs1 and mcs2 of pVIVO2-mcs respectively. The expression of fusion antigen protein in MCF-7 cell line transfected with pVIVO2-SjFABP/Sj26.GAPDH was determined by indirect immunofluorescence assay (IIF) and reverse transcriptase-polymerase chain reaction (RT-PCR). The evaluation of vaccine’s immuno-protection efficacy is tested by pharmacological trials. Results: The trivalent-epitope DNA vaccine pVIVO2-SjFABP/Sj26.SjGAPDH was successfully constructed. In the trivalent-epitope DNA vaccine group, the worm reduction rate and the egg reduction rate were 58.6% and 59.8% compared with control group. Conclusion: The trivalent-epitope DNA vaccine pVIVO2-SjFABP/Sj26.SjGAPDH could induce higher protection efficiency than both monovalent and bivalent vaccine, and paves the way for development of DNA vaccine against Schistosoma japanicum.
Key words: Schistosama japanicum    SjFABP    SjGAPDH    Sj26    Trivalent-epitope DNA vaccine
收稿日期: 2009-12-30 出版日期: 2010-05-25
基金资助:

国家“863”计划（2004AA2Z3212）、教育部博士点基金（20070487059）、教育部新世纪优秀人才计划（NCET060646）资助项目
通讯作者: 余龙江     E-mail: yulj@mail.hust.edu.cn
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引用本文:

朱路 龙全科 刘海峰 鲁明波 胡媛 石佑恩 余龙江. 日本血吸虫三价DNA疫苗pVIVO2SjFABP/Sj26.SjGAPDH的构建及其免疫保护作用评价[J]. 中国生物工程杂志, 2010, 30(05): 36-42.

SHU Lu, LONG Quan-Ke, LIU Hai-Feng, LU Meng-Bei, HU Yuan, DAN You-En, TU Long-Jiang. Construction and Immuno-protection Evaluation of Trivalent-epitope DNA Vaccine pVIVO2-SjFABP/Sj26.SjGAPDH. China Biotechnology, 2010, 30(05): 36-42.

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https://manu60.magtech.com.cn/biotech/CN/    或    https://manu60.magtech.com.cn/biotech/CN/Y2010/V30/I05/36

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