猪Ghrelin与IGF-1融合基因克隆和表达研究

Study on Cloning and Expression of Porcine Ghrelin and IGF-1 Fusion Gene

目的本试验克隆和构建了猪Ghrelin(GL)与类胰岛素生长因子-1(IGF-1)的融合基因及其表达载体,研究它们在体外HEK293细胞的表达,为探索新型高效的动物生长和免疫调节生物制剂奠定了基础。方法利用基因重叠延伸PCR技术分别获得了GL、短链IGF-1基因及其融合基因(GI),双酶切后插入到VR1020真核分泌型表达载体上,分别命名为VGL、VRI和VGI。用壳聚糖(Chitosan,CS)和mPEG-PEI-CS分子包裹VRI、VGH和VGI制备纳米颗粒,先后进行HEK293细胞转染表达实验。结果成功克隆了猪Ghrelin基因及其与IGF-1的融合基因,构建了它们的真核表达载体,并进行了纳米分子包装,在HEK293细胞获得高效表达。结论猪Ghrelin和IGF-1融合基因及其真核载体的成功构建和表达,为进一步研发安全有效的新型生物制剂、促进动物的生长发育和免疫机能提供了新途径。

Objective To develop safe and effective novel regulator for animal growth and immunity, the expression vector containing pig ghreliu （GL） and IGF-1 （GI） genes were constructed and expressed in HEK293 cells. Methods Pig ghrelin （GL） gene and its fusion gene IGF-1 （GI） were constructed and digested by BamHI and BglII restriction enzyme. Then the segments were cloned into the VR1020 eukaryotic vector and named VGL, VR1, VGI, respectively, Chitosan （CS） and its derivative （mPEG-PEI-CS） were chosen to pack VRI, VGL and VGI to prepare nanoparticles for transfection of HEK293 cell. Results The results showed that the recombinant eukaryotic expression plasmid containing GL and IGF-1 （GI） genes was successfully constructed and encapsulated with chitosan and mPEG-PEI-CS modified molecule. Finally, these genes were effectively expressed in HEK293 ceils. Conclusion The successful construction and expression of pig ghrelin and IGF-1 fusion gene would inspire and facilitate further development of safe and efficacious regulator for animal growth and im-munity.