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浙江大学学报（农业与生命科学版）
Journal of Zhejiang University: Agric. & Life Sci.  2010, Vol. 36 Issue (6): 609-614    DOI: 10.3785/j.issn.1008-9209.2010.06.003
Biological sciences & biotechnology     
Expression of Acc-royalisin from royal jelly of Apis cerana cerana in Escherichia coli and preparation of its polyclonal antibody
DING Mei-hui,JIN Feng,SHEN Li-rong,CHEN Zheng-xian
DING Mei-hui,School of Biosystems Engineering and Food Science,Zhejiang University,Hangzhou310029,China JIN Feng,School of Biosystems Engineering and Food Science,Zhejiang University,Hangzhou310029,China SHEN Li-rong,School of Biosystems Engineering and Food Science,Zhejiang University,Hangzhou310029,China CHEN Zheng-xian,School of Biosystems Engineering and Food Science,Zhejiang University,Hangzhou310029,China
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Abstract  The coding region of pre-pro-Acc-royalisin was amplified by PCR from cDNA library of the Chinese honeybee,Apiscerana cerana head,and was cloned into the vector pGEX-4T-2 for expression in Escherichia coli BL21 . The expressed fusion protein,glutathione S-transferase (GST)-pre-pro-Acc-royalisin of36 ku was obtained,which was cross-reacted with GST antibody accounting for up to16 .3% of bacterial protein . With the expressed products retrieved from the SDS-PAGE gels as antigen to immunize New Zealand white rabbits,the polyclonal antibody was prepared .With the purified recombinant GST-pre-pro-Acc-royalisin fusion protein as antigen,the high titers of the antibody was shown with ELISA analysis . The specificity of the antibody against the same antigens was then confirmed by Western blot .This study provides a new tool for the detection of antimicrobial of royal jelly,biological product quality of royalisin and resistance of honeybee .

Published: 09 November 2010
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DING Mei-hui
JIN Feng
SHEN Li-rong
CHEN Zheng-xian
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DING Mei-hui,JIN Feng,SHEN Li-rong,CHEN Zheng-xian. Expression of Acc-royalisin from royal jelly of Apis cerana cerana in Escherichia coli and preparation of its polyclonal antibody. Journal of Zhejiang University: Agric. & Life Sci., 2010, 36(6): 609-614.

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http://www.zjujournals.com/agr/10.3785/j.issn.1008-9209.2010.06.003     OR     http://www.zjujournals.com/agr/Y2010/V36/I6/609


中华蜜蜂王浆抗菌肽Acc- royalisin基因原核表达及多克隆抗体制备

以含中华蜜蜂王浆抗菌肽 Acc-royalisin基因的该蜂头部 cDNA文库质粒为模板,用 PCR方法扩 增出其前体片段,并克隆入表达载体 pGEX-4T-2中,在大肠杆菌 BL21中与谷胱甘肽 S-转移酶(GST)融合 进行表达,得到大小为36 ku,占细胞总蛋白16 .3%,与 GST多克隆抗体有免疫反应的表达产物 ;用割胶 回收获得的目的蛋白注射新西兰大白兔,采集血清制备多克隆抗体,该抗体与纯化的 GST-Acc-royalisin 融合蛋白经 ELISA反应显示出较高效价,经 Western blot印迹分析获得特异性条带 。这为进一步利用 免疫方法检测王浆的防腐性能和 Acc-royalisin生物制品质量,以及蜜蜂抗病性奠定了基础 。
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