鸡B-F2基因真核表达系统的建立与鉴定  

Construction and identification of eukaryotic expression system of chicken B-F2 gene

作　　者：金园园[1] 陈芳芳[1] 余为一[1] 

机构地区：[1]安徽农业大学,安徽省人兽共患病重点实验室,安徽合肥230036

出　　处：《畜牧与兽医》2012年第9期1-3,共3页Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金(No.31172306)资助

摘　　要：鸡B-F2基因编码的主要组织相容性复合体(MHC)I类分子α链在递呈内源性抗原中起重要作用。本研究应用RT-PCR方法从鸡外周血单核细胞中扩增了B-F2基因片段,其大小为1 068 bp。进一步将其定向插入真核表达载体,构建重组质粒pEGFP-N1-B-F2。经脂质体法转染COS7,所表达的α链定位于在细胞的浆膜,同时转染P815细胞,经G418的筛选(0.6 mg/mL)得到稳定表达细胞株。该细胞株经培养10代次后,仍在RT-PCR中能获得相应DNA片段。上述结果表明,B-F2基因在动物真核表达系统得到表达,建立的P815(B-F2)细胞株为进一步研究MHC I类分子α链在免疫应答中作用奠定了基础。B-F2 gene, which encodes a chain of chicken major histocompatibility complex （MHC） class I molecule, plays an important role in endogenous antigen presentation. In this study, the B-F2 gene fragment with a length of 1068bp was amplified by RT-PCR method from chicken peripheral blood mononuclear cell （PBMC）. Then this fragment was inserted into pEGFP-N1, and thus the vector pEGFP-N1- B-F2 was constructed. Further the recombinant plasmid was transfected into COS＇/with Lipofectin reagent. The B-F2 gene product was loca- ted to plasma membrane of the cells; meanwhile it was transfected into P815 cells and a positive strain, P815 （B-F2）, was selected by G418 （0. 6mg/mL）. After the transfected cells were passaged 10 times, the specific B-F2 fragment could be amplified by RT-PCR from the cells. All these results showed that the eukaryotic system with stable expression of B-F2 was successfully established. This result provided a founda- tion for further study on the roles of MHC I class molecule o~ chain in immune response.

关 键 词：B-F2 鸡 主要组织相容性复合体(MHC) 真核表达系统 

分 类 号：S852.4[农业科学—基础兽医学]