| J亚群禽白血病病毒gp85基因在毕赤酵母中的分泌表达 |
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| 引用本文： | 徐延伟,高宏雷,高玉龙,李凯,高立,秦立廷,祁小乐,王永强,王笑梅.J亚群禽白血病病毒gp85基因在毕赤酵母中的分泌表达[J].中国畜牧兽医,2012,39(4):42-46. |
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| 作者姓名： | 徐延伟 高宏雷 高玉龙 李凯 高立 秦立廷 祁小乐 王永强 王笑梅 |
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| 作者单位： | 中国农业科学院哈尔滨兽医研究所禽传染病研究室,兽医生物技术国家重点实验室,黑龙江哈尔滨 150001 |
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| 基金项目： | 现代农业肉鸡产业技术体系建设(nycyts-42-G3-01) |
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| 摘 要： | 为获得J亚群禽白血病病毒gp85蛋白,将gp85基因克隆至毕赤酵母表达载体pPIC9K,构建了pPIC9K-gp85表达载体。将鉴定正确的表达载体经Sal Ⅰ酶切线性化后电转化毕赤酵母宿主菌GS115,经MD及G418抗性平板筛选后挑取阳性克隆进行PCR鉴定,获得重组酵母菌株GS115/pPIC9K-gp85。对阳性重组菌进行诱导表达,表达产物经SDS-PAGE、Western blotting和Dot-ELISA分析,结果表明,gp85基因在毕赤酵母中获得分泌表达,表达产物与J亚群禽白血病病毒多克隆抗体有良好的反应活性,分泌表达量达40 mg/L。本研究为J亚群禽白血病病毒诊断方法的建立及gp85蛋白的深入研究奠定了基础。
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| 关 键 词： | J亚群禽白血病 gp85基因 毕赤酵母 分泌表达 |
| 收稿时间： | 2011-11-05 |
Secretory Expression of Avian Leukosis Virus Subgroup J gp85 Gene in Pichia pastoris |
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| XU Yan-wei
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GAO Hong-lei
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GAO Yu-long
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LI Kai
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GAO Li
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QIN Li-ting
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QI Xiao-le
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WANG Yong-qiang
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WANG Xiao-mei.Secretory Expression of Avian Leukosis Virus Subgroup J gp85 Gene in Pichia pastoris[J].China Animal Husbandry & Veterinary Medicine,2012,39(4):42-46. |
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| Authors: | XU Yan-wei GAO Hong-lei GAO Yu-long LI Kai GAO Li QIN Li-ting QI Xiao-le WANG Yong-qiang WANG Xiao-mei |
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| Institution: | Division of Avian Infectious Diseases,State Key Laboratory of Veterinary Biotechnology,Harbin Veterinary Research Institute, Chinese Academy Agricultural Sciences,Harbin 150001,China |
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| Abstract: | To express recombinant gp85 protein of avian leukosis virus subgroup J(ALV-J) in P.pastoris,the gp85 gene was amplified and cloned into the pPIC9K vector.The recombinant plasmid pPIC9K-gp85 was identified,linearized with Sal Ⅰ and transformed into P.pastoris strain GS115 by electroporation.Transformants were selected on MD plates then on YPD plates containing G418 and confirmed by PCR.SDS-PAGE assay demonstrated that gp85 protein was expressed and secreted into the culture medium.The expression product amounted to 40 mg/L of culture under optimized condition.The recombinant gp85 protein showed good antigenicity in Western blotting and Dot-ELISA assays.This study laid the foundation for further study of the gp85 protein and the development of the diagnostic methods for ALV-J. |
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| Keywords: | ALV-J gp85 gene Pichia pastoris secretory expression |
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