家蚕B类清道夫受体BmSCRB8基因的克隆及表达

赵羽卒1 张奎1 唐梅1 徐曼1 李重阳1 潘光照1 申利1 崔红娟1 杨丽群1

(1.西南大学家蚕基因组生物学国家重点实验室, 重庆 400716)

【摘要】B类清道夫受体在动脉粥样硬化及其他心血管疾病的形成或抑制,机体免疫防御,凋亡细胞清除等生理过程中起着重要的作用。克隆了家蚕B型清道夫受体家族的一个成员BmSCRB8基因,通过RACE技术获得BmSCRB8的c DNA全长为2 668 bp,其ORF为1 704 bp,编码567个氨基酸,通过在线预测其蛋白分子量为63.87 k Da,等电点为6.06。采用RT-PCR方法得到了BmSCRB8的时空表达谱,结果表明BmSCRB8在家蚕各组织以及血液各时期均有表达,且在脂肪体中表达量最高,变态发育时期表达量较高。原核表达获得BmSCRB8重组蛋白,并经由蛋白纯化、免疫小鼠后制备得到家蚕BmSCRB8多克隆抗体。同时构建了BmSCRB8真核表达载体并转染家蚕胚胎细胞系。免疫荧光及过表达结果显示BmSCRB8主要定位于细胞膜上,Western blotting结果显示免疫小鼠后所得到的抗血清可特异性识别BmSCRB8蛋白。

【关键词】 家蚕; BmSCRB8; 表达谱; 多克隆抗体; 亚细胞定位;

【DOI】

【基金资助】 国家重点基础研究发展计划(973计划)(No.2012CB114603) National Basic Research Program of China(973 Program)(No.2012CB114603) 国家自然科学基金(No.81201551) National Natural Science Foundation of China(No.81201551) 中央高校基本科研业务费专项资金(No.XDJK2013B020) the Fundamental Research Funds for the Central Universities(No.XDJK2013B020) 中央高校基本科研业务费专项资金学科团队项目(No.2362015XK09) Disciplinary Team Program of the Fundamental Research Funds for the Central Universities(No.2362015XK09) 重庆市研究生科研创新项目(No.CYB2015067) Postgraduate Technology Innovation Program of Chongqing(No.CYB2015067) 西南大学基本科研业务费专项资金(No.XDJK2015D021)资助 Basic Research Specialized Funds of Southwest University(No.XDJK2015D021)

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This Article

ISSN:1000-3061

CN: 11-1998/Q

Vol 32, No. 10, Pages 1408-1421

October 2016

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摘要

  • 1 材料与方法
  • 2 结果与分析
  • 3 讨论
  • 参考文献