长角血蜱卵泡抑素相关蛋白的定性-微生物学报.pdfVIP

生 物 工 程 学 报 Chin J Biotech 2009, November 25; 25(11): 1646-1651 Chinese Journal of Biotechnology ISSN 1000-3061 cjb@ © 2009 Institute of Microbiology, CAS CSM, All rights reserved 动物及兽医生物技术 田占成, 刘光远, 殷宏, 罗建勋, 谢俊仁 , 730046 : 参照GenBank 中长角血蜱致病性Okayama 株卵泡抑素基因的核苷酸序列(GenBank Accession No. DQ248886) 设计合成一对引物, 从本实验室保藏的单克隆洁净长角血蜱饥饿成蜱中快速提取总 RNA, 通过 RT-PCR 扩增出 814 bp 的卵泡抑素基因, 序列比对结果显示:与长角血蜱致病性Okayama 株的核苷酸序列及氨基酸序列一致性分别为 97.8%和 99%, 将其亚克隆到表达载体 pGEX-4T-1 中进行表达, GST 融合重组蛋白预期分子量为 57 kD 。表达重组蛋白经 MagneGSTTM 蛋白纯化系统纯化后作为抗原分别与抗不同发育阶段长角血蜱( 卵、幼蜱、若蜱、成蜱) 多克隆抗体作为一 抗进行免疫印迹, 结果表明: 与长角血蜱卵制备的多克隆抗体有很强的免疫反应, 而与其他发育阶段(幼蜱、若蜱、成蜱) 饥饿长角血蜱制备的多克隆抗体反应性很弱。以上结果表明: 长角血蜱卵泡抑素蛋白在长角血蜱产卵及卵成熟发育时期 的表达水平较其他发育阶段(幼蜱、若蜱、成蜱) 的蛋白表达水平高。 : 长角血蜱, 卵泡抑素, 定性 Characterization of follistatin-related protein from the hard tick Haemaphysalis longicornis Zhancheng Tian, Guangyuan Liu, Hong Yin, Jianxun Luo, and Junren Xie Key Laboratory of Veterinary Parasitology of Gansu Province, State Key Laboratory of Veterinary Etiological Biology, Lanzhou Veterinary Research Institute , Chinese Academy of Agricultural Sciences, Lanzhou 730046, China Abstract: We designed the primers based on the sequence of the follistatin-related protein from Haemaphysalis longicornis Okayama strain accessed in GenBank. We cloned a gene encoding follistatin-related protein by RT-PCR, and the length cDNA is 814 bp, encoding a deduced protein of 289 amino acids. The alignment with the sequence of follist