鸡IL-2和IL-4融合基因真核表达质粒的构建及其表达产物的生物学活性  

Construction of Eukaryotic Expression Plasmid of Chicken IL-2 and IL-4 Fusion Gene and Biological Activity of its Expression Products

作　　者：徐彤[1] 贡鑫 郑明学[1] 吕晓玲 郑龙龙 崔凯玲 张雪松 段步婷 白瑞[1] XU Tong;GONG Xin;ZHENG Ming-xue;LYU Xiao-ling;ZHENG Long-long;CUI Kai-ling;ZHANG Xue-song;DUAN Bu-ting;BAI Rui(College of Veterinary Medicine,Shanxi Agricultural University,Taigu 030801,China)

机构地区：[1]山西农业大学动物医学学院,山西太谷030801

出　　处：《中国兽医杂志》2022年第4期30-34,共5页Chinese Journal of Veterinary Medicine

基　　金：国家自然科学基金项目(31972647)。

摘　　要：为提高鸡球虫病活疫苗的免疫效果并降低其副作用,本试验用合成的鸡白介素2(chIL-2)和鸡白介素4(chIL-4)基因片段构建3个融合基因真核表达质粒,并利用脂质体法将其转染至原代鸡胚盲肠上皮细胞,通过实时荧光定量PCR(RT-qPCR)检测其表达水平,MTT法测定其表达的分泌产物活性。结果显示:成功构建的pCI-chIL-4-chIL-2-EGFP、pCI-chIL-2-EGFP、pCI-chIL-4-EGFP重组真核表达质粒能在原代鸡胚盲肠上皮细胞中高效表达;3种重组真核表达质粒转染后24~96 h的鸡脾淋巴细胞活性极显著高于空载组(P<0.01),且在72 h时达峰值;而单一因子组鸡脾淋巴细胞增殖活性极显著低于pCI-chIL-4-chIL-2-EGFP组(P<0.01)。由此可见,本试验成功构建了能表达分泌chIL-2和chIL-4的融合基因重组质粒;且chIL-4-chIL-2融合蛋白生物学活性显著高于单一因子。In order to improve the immune effect and reduce the side effects of avian coccidiosis vaccine,three eukaryotic expression vectors of fusion gene were constructed with the synthetic chicken interleukin 2(chIL-2)and chicken interleukin 4(chIL-4)gene fragments;transfected via liposome method into primary chicken embryo cecal epithelial cells,their expression levels were assessed by real-time fluorescence quantitative PCR,and the activity of their expression and secretion products was determined by MTT method.The results showed that the recombinant eukaryotic expression plasmids pCI-chIL-4-chIL-2-EGFP,pCI-chIL-2-EGFP and pCI-chIL-4-EGFP were successfully constructed and expressed in primary cecal epithelial cells of chicken embryo.The spleen lymphocyte activity of three recombinant eukaryotic expression plasmids was significantly higher than that of the control group(P<0.01)24-96 h after transfection,and reached the peak at 72 h.The proliferation activity of spleen lymphocytes in the single factor group was significantly lower than that of PCI-chIL-4-chIL-2-EGFP group(P<0.01).Thus,this study successfully constructed recombinant plasmids expressing chIL-2 and chIL-4 fusion protein,and the biological activity of chIL-4-chIL-2 fusion protein was significantly higher than that of single factor.

关 键 词：细胞因子 鸡白介素2 鸡白介素4 生物学活性 基因佐剂 

分 类 号：S855.9[农业科学—临床兽医学]