(猪带绦虫六钩蚴TSOL16基因的克隆.docVIP

(猪带绦虫六钩蚴TSOL16基因的克隆分析 1,2，，2，才学鹏1中国农业科学院兰州兽医研究所 家畜疫病病原生物学国家重点实验室 甘肃省动物寄生虫病重点实验室甘肃 兰州 7300462．新疆农业大学 动物学院，830052） 摘要：用RT-PCR技术扩增基因将扩增产物与MD-18T载体连接，重组PCR、酶切鉴定后进行测序pGEX-4T-1原核表达载体，用IPTG诱导表达后进行SDS、Western-blot分析；将所表达的蛋白免疫小鼠，经ELISA检测血清抗体，验证其免疫原性。结果显示：所克隆的基因片段长度为bp（碱基），含有1个bp的开放阅读框架，编码个氨基酸与已报道的基因核苷酸序列同源性为免疫小鼠在周后即检测到血清抗体，30天即达到较高水平说明该融合蛋白具有较好的免疫原性。 关键词基因克隆loning expression and immunogenicity study on TSOL16 gene from Taenia solium oncosphere MA Quan-ying1,2,LUO Xue-nong1,JING Zhi-zhong1,WANG Ying2,YUE Cheng2,CAIXue-peng1** (1.Key Laboratory of Animal Parasitology of Gansu Province, State Key Laboratory of Veterinary Etiological Biology ,Lanzhou Veterinary Research Institute，Chinese Academy of Agricultural Sciences, Lanzhou 730046 ,China;.College of Veterinary Medicine, Xinjiang agricultural University, Urumqi 830052 China;) Abstract：The T immunogen gene of Taenia solium oncosphere was amplified by RT-PCR, and ligated into pMD-18T vector. The recombinant plasmid was certified by PCR amplification, digestion with endonuclease and sequencing. A prokaryotic expression plasmid pGEX-TSOL16 was constructed by inserting the TSOL16 gene into pGEX-4T-1. The expression was induced by IPTG, the cultures were collected at different times, and the expressed protein was analyzed by SDSand Western-blotting. The mice were immunized with the fusion protein, and antibody of the TSOL16 protein was tested by ELISA. Results showed that the TSOL16 cDNA was 559 bp in length and had an ORF which was 402 bp in length and encoded 134 amino acids. Homology of nucleotide sequence between the cloned TSOL16 gene and the sequence from GenBank was 99.2 ％.Key words: Taenia solium oncosphere; TSOL16 gene; clone; expression ; immunogenicit 猪囊尾蚴（Cysticercus celluosae)病是一种由猪带绦虫（Taenia soJium）幼虫引起的严重危害畜牧业生产和人类身体健康的人畜共患寄生虫病，是目前世界范围内公认的社会、政治、经济病[1，2]。 对于囊尾蚴病的免疫预防研究，开展较早的有全囊虫匀浆抗原[3]、六钩蚴抗原[4]、六钩蚴排泄／分泌抗原、囊尾蚴体外培养的分泌代谢抗原和组织培养的细胞抗原等疫苗的研究，并且已用于免疫保护性试验[5]。结果表明，这些抗原作为疫苗均能取得良好的预防效果，可使免疫动物获得抗虫卵攻击感染的坚强抵抗力，多数动物能得到完全保护。但是，这些