鸡白细胞介素-2原核表达及其活性鉴定  

Prokaryotic Expression and Activity Identification of Chicken Interleukin-2

作　　者：韩顺子 黄霞 孟闯[1,2,3] 耿士忠 康喜龙[1,2,3] 潘志明 焦新安[1,2,3] HAN Shunzi;HUANG Xia;MENG Chuang;GENG Shizhong;KANG Xilong;PAN Zhiming;JIAO Xinan(Jiangsu Key Laboratory of Zoonoses,Yangzhou University,Yangzhou,Jiangsu 225009;Key Laboratory of Prevention and Control of Biological Hazard Factors(Animal Origin)for Agrifood Safety and Quality of Ministry of Agriculture and Rural Affairs,Yangzhou University,Yangzhou,Jiangsu 225009;Jiangsu Co-innovation Center for Prevention and Control of Important Animal Infectious Diseases and Zoonoses,Yangzhou,Jiangsu 225009)

机构地区：[1]扬州大学江苏省人兽共患病学重点实验室,江苏扬州225009 [2]扬州大学农业农村部农产品质量安全生物性危害因子(动物源)控制重点实验室,江苏扬州225009 [3]江苏高校动物重要疫病与人兽共患病防控协同创新中心,江苏扬州225009

出　　处：《中国家禽》2024年第5期56-61,共6页China Poultry

基　　金：江苏省重点研发计划(现代农业)项目(BE2021331、BE2021354);江苏省政策引导类计划(国际科技合作港澳台科技合作)资助项目(BZ2020013)。

摘　　要：试验旨在获得具有生物活性的鸡白细胞介素-2(ChIL-2)重组蛋白。试验以克隆获得的ChIL-2基因为模板,分别构建原核重组表达菌BL21(pCold-ChIL-2)和BL21(pGEX-6P-1-ChIL-2),进行诱导表达和纯化;通过BrdU ELISA法检测重组蛋白诱导鸡脾脏淋巴细胞的增殖情况,评估其生物活性;以rHis-ChIL-2免疫小鼠制备多克隆抗体。结果显示:成功构建了原核重组表达菌BL21(pCold-ChIL-2)、BL21(pGEX-6P-1-ChIL-2);经表达纯化后分别获得纯度较高的rHis-ChIL-2和rGST-ChIL-2蛋白,大小分别为14 ku和39.4 ku;rHis-ChIL-2和rGST-ChIL-2蛋白分别能够促进脾脏淋巴细胞显著或极显著增殖(P<0.01或P<0.001),均具有良好的生物活性;以纯化的rHis-ChIL-2免疫小鼠制备的多克隆抗体效价为5.12×105,可识别rHis-ChIL-2和rGST-ChIL-2,与真核表达的ChIL-2蛋白反应。结果表明,原核表达的重组蛋白ChIL-2具有良好的生物活性,采用其制备的多克隆抗体可识别真核表达的ChIL-2蛋白,为后期ChIL-2的检测奠定基础。The experiment was aimed to obtain recombinant chicken interleukin-2(ChIL-2)with biological activity.The experiment used the ChIL-2 gene obtained by cloning as a template,and constructed the prokaryotic recombinant expression bacte-ria BL21(pCold-ChIL-2)and BL21(pGEX-6P-1-ChIL-2)for induced expression and purification respectively.The recombinant ChIL-2 was expressed and purified.The proliferation of chicken spleen lymphocytes induced by the recombinant protein ChIL-2 was detected using BrdU ELISA.rHis-ChIL-2 was used as an immunogen to produce polyclonal antibodies.The results showed that the prokaryotic recombinant expression bacterium BL21(pCold-ChIL-2)and BL21(pGEX-6P-1-ChIL-2)were successfully constructed.rHis-ChIL-2 and rGST-ChIL-2 proteins with high purity were obtained,with sizes of 14 ku and 39.4 ku,respectively.rHis-ChIL-2 and rGST-ChIL-2 could significantly promote the proliferation of splenic lymphocytes(P<0.01 or P<0.001).The titer of polyclonal antibodies was 5.12×105,which could recognize the recombinant proteins rHis-ChIL-2 and rGST-ChIL-2,and react with the eukaryotic expressed ChIL-2 protein.The results suggested that the prokaryotic expressed recombinant protein ChIL-2 had good biological activity and the prepared ChIL-2 polyclonal antibodies could recognize the eukaryotic expressed ChIL-2 protein,which laid the foundation for the detection of ChIL-2.

关 键 词：ChIL-2 原核表达 生物活性 多克隆抗体 

分 类 号：S855.3[农业科学—临床兽医学]