中国兽医科学2011，Vol.41Issue(5)：519-525,7.
隐孢子虫鼠基因型CP15基因真核表达质粒的构建及在Hela细胞中的表达
Construction of eukaryotic expression plasmids with Cryptosporidium mouse genotype CP15 gene and their expression in Hela cells
摘要
Abstract
In order to construct recombinant expression plasmids with Cryptosporidium mouse genotype CP15 gene and express them in Hela cell,CP15 gene and CP15 gene containing CpG-ODN(C-CP15) were amplified from pMD18-T-CP15 by PCR. The CP15 gene and C-CP15 gene were identified by enzyme digestion and sequencing. Then the CP15 gene and C-CP15 gene were cloned into the pVAX 1 vector. Then Hela cells were transfected with recombinant expression plasmids by Lipofectarnine and assayed by RTPCR, indirect immunofluorescence, SDS-PAGE and Western-blot assay. In results, the CP15 gene was about 360 bp and the C-CP15 gene was about 380 bp in size. The recombinant plasmids pVAX1-CP15 and pVAX1C-CP15 were constructed successfully,and confirmed by enzyme digestion and sequencing. RT-PCR,indiret immunofluorescence, SDS-PAGE and Western-blot assay showed that the recombinant plasmids were effectively transcripted and expressed in Hela cell, and the expressed product had good immunity activity. The results provide the basis for further research and development of higher protective vaccines against cryptosporidiosis.关键词
隐孢子虫鼠基因型/CP15/真核表达质粒/Hela细胞/表达Key words
Cryptosporidium mouse genotype/ CP15/ eukaryotic expression plasmid/ Hela cell/ expression分类
农业科技引用本文复制引用
苏庆美,何生虎,米荣升,张国恩,黄燕,周鹏,秦培兰,呼高伟,陈兆国..隐孢子虫鼠基因型CP15基因真核表达质粒的构建及在Hela细胞中的表达[J].中国兽医科学,2011,41(5):519-525,7.基金项目
国家"十一五"高技术研究发展计划(863)项目(2006AA10A207) （863）
国家"十一五"科技支撑计划项目(2007BAD40B05) （2007BAD40B05）
上海市科技兴农重点攻关项目[沪农科攻字(2005)第3-4号] （2005）
中央级公益性科研院所基本科研业务费专项资金项目(2010JB12) （2010JB12）
