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堆型艾美球虫表面抗原基因 cSZ1 在大肠埃希氏菌中的表达
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覃宗华1， 蔡建平1， 谢明权1， 艾哈迈德2， 彭新宇1， 魏文康1，
吴惠贤1
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(1. 广东省农业科学院 兽医研究所， 广东 广州510640；
2. 尼亚拉大学 兽医学院， 苏丹 尼亚拉)
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摘要：
根据已克隆的堆型艾美球虫( Eimeria acervulina )广东株子孢子表面抗原基因cSZ1的cDNA序列设计了特异性引物，用PCR方法扩增cSZ1的开放阅读框架（ORF）后克隆至表达载体pET32a(+)，构建了重组表达质粒pET32a(+)cSZ1，并将其转化至大肠埃希氏菌BL21(DE3)。经IPTG诱导，获得了cSZ1重组抗原在大肠埃希氏菌中的高效表达，表达产物量可达菌体总蛋白的9.3%，融合蛋白的分子质量约为40ku。重组菌诱导表达的产物经SDSPAGE后，用堆形艾美球虫感染鸡的超免疫血清进行免疫印迹分析，结果为阴性，提示cSZ1所编码的抗原可能主要是T细胞抗原表位。 关键词：
艾美球虫； 大肠埃希氏菌； 表达
中图分类号：S 852.723 文献标识码：
A 文章编号：1000-6419(2004)08-0022-05 |
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Expression of sporozoite surface antigenic gene cSZ1 of
Eimeria acervulina Guangdong strain in Escherichia coli
QIN Zonghua1， CAI Jianping1， XIE
Mingquan1， Ahmed Bashar Elhag2， PENG
Xinyu1， WEI Wenkang1， WU Huixian1
(1. Institute of Veterinary Medicine, Guangdong Academy of Agricultural Sciences, Guangzhou
510640,China； 2. Faculty of Veterinary Science, University of Nyala， Nyala,Sudan) |
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Abstract: The ORF of cSZ1 gene of Eimeria acervulina Guangdong strain was amplified by PCR using the specific primers designed according to the sequences of cSZ1(Gd) gene cloned in our laboratory, and its ligated products with vector pET32a(+) were transformed into E.coli BL21(DE3) by the
CaCl2 method. The transformants were identified by PCR amplification and endonuclease digestion. The sequences of positive clone were analyzed,and the
recombinant protein was induced to be expressed by 1mmol/L IPTG in vitro . The molecular weight of cSZ1 recombinantprotein is about
40ku, and the amount of recombinantprotein in total bacteria protein wasabout 9.3% . Western blotting result of the purified cSZ1recombinantprotein
was negativewhen chicken hyperimmune serum of E. acervulina was used as a probe.
Key words: Eimeria acervulina ； Escherichia coli ； expression |
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