环形泰勒虫表面抗原重组蛋白Tasp-Tams1-Spag1生物信息学分析与原核表达
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摘要
运用生物信息学软件对环形泰勒虫表面抗原串联基因Tasp-Tams1-Spag1进行分析,预测其编码蛋白的主要特性与抗原表位等,并对此串联基因进行克隆与原核表达。结果表明:串联重组蛋白Tasp-Tams1-Spag1属于不稳定非分泌型亲水性蛋白;编码346个氨基酸;有4个低复杂性的结构域,且含有Sorb与Btz、NL、NOT的同源区域;可能存在11个B细胞表位优势区段与17个T细胞表位优势区段,含有T、B细胞联合表位,表明Tasp-Tams1-Spag1重组蛋白可能具有良好的免疫原性。IPTG诱导重组蛋白原核表达,结果显示,该重组蛋白以包涵体形式存在;经Western blot检测,表明此串联蛋白反应原性良好,为后续深入研究免疫抗原奠定基础。
The main characters and antigenic epitopes of the recombinant protein Tasp-tams1-spag of Theileria annulata were analyzed and predicted in the present study by using the biological software and online software in order to clone the surface antigen Tasp-tams 1-spag gene and to construct prokaryotic expression vectors. The recombinant Tasp-tams1-spag contained 346 amino acids and belonged to unstable hydrophilic non-secreted protein. It contained 4 low complexity domains as well as Sorb, Btz, NL and NOT homologous regions. There might be 11 B-cell major epitope domains, 17 T-cell major epitope domains and two cross-reactive epitopes. The recombinant protein was predicted to have good immunogenicity in theory. Subsequently, the recombinant Tasp-tams1-spag was expressed by induction with IPTG and expression condition was optimized. The expressed protein was mainly obtained in the form of inclusion bodies. Western blot assay also revealed that the recombinant protein had good antigenicity.
The main characters and antigenic epitopes of the recombinant protein Tasp-tams1-spag of Theileria annulata were analyzed and predicted in the present study by using the biological software and online software in order to clone the surface antigen Tasp-tams 1-spag gene and to construct prokaryotic expression vectors. The recombinant Tasp-tams1-spag contained 346 amino acids and belonged to unstable hydrophilic non-secreted protein. It contained 4 low complexity domains as well as Sorb, Btz, NL and NOT homologous regions. There might be 11 B-cell major epitope domains, 17 T-cell major epitope domains and two cross-reactive epitopes. The recombinant protein was predicted to have good immunogenicity in theory. Subsequently, the recombinant Tasp-tams1-spag was expressed by induction with IPTG and expression condition was optimized. The expressed protein was mainly obtained in the form of inclusion bodies. Western blot assay also revealed that the recombinant protein had good antigenicity.
引文
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[12]曹雯丽,陈亮,刘启生,等.新疆牛环形泰勒虫病的流行现状和防治[J].草食家畜,2011,(3):76-78.
[2]汪明.兽医寄生虫学[M].3版.北京:中国农业大学出版社,2008:332.
[3]农业部畜牧兽医局.一、二、三类动物疫病释义[M].北京:中国农业出版社,2004:97-101.
[4]蔺红玲,张继瑜,魏小娟,等.环形泰勒虫主要膜蛋白的研究进展[J].黑龙江畜牧兽医,2011,(5):22-24.
[5]张艳贞,宣劲松.蛋白质科学-理论、技术与应用[M].北京:北京大学出版社,2012,11-152.
[6]黄家雨.新疆牛环形泰勒虫Tams1基因的克隆、表达与巢式PCR诊断方法的建立[D].新疆:新疆农业大学,2008.
[7]Ghoneim A M,EI-Fayomy A O.Targeting tams-1 gene results in underestimation of Theileria annulata infection in diseased cattle in Egypt[J].Acta Parasitol,2014,59(1):85-90.
[8]Darghouth M A,Boulter N R,Gharbi M,et al.Vaccination of calves with an attenuated cell line of Theileria annulata and the sporozoite antigen SPAG-1 produces a synergistic effect[J].Vet Parasitol,2006,142(1-2):54-62.
[9]Salih D E A,Ahmed J S,Bakheit M A,et al.Validation of the indirect Ta SP enzyme-linked immunosorbent assay for diagnosis of Theileria annulata infection in cattle[J].Parasitol Res,2005,97(4):302-308.
[10]Bakheit M A,Schnittger L,Salih D A,et al.Application of the recombinant Theileria annulata surface protein in an indirect ELISA for the diagnosis of tropical theileriosis[J].Parasitol Res,2004,92(4):299-302.
[11]Mohamed A M,Abdel-Rady A,Ahmed L S,et al.Evaluation of indirect Ta SP enzyme-linked immunosorbent assay for diagnosis of tropical theileriosis in cattle(Bos indicus)and water buffaloes(Bubalus bubalis)in Egypt[J].Vet Parasitol,2012,186(3-4):486-489.
[12]曹雯丽,陈亮,刘启生,等.新疆牛环形泰勒虫病的流行现状和防治[J].草食家畜,2011,(3):76-78.