人TSHR289基因重组腺病毒的制备与鉴定  

Preparation and identification of recombinant adenovirus containing human TSHR289 gene

作　　者：周军[1] 马洁[1] 汤新逸[1] 田洁[1] 许化溪[1] 王胜军[1] ZHOU Jun;MA Jie;TANG Xin-yi;TIAN Jie;XU Hua-xi;WANG Sheng-jun(Institute of Laboratory Medicine, School of Medicine, Jiangsu University, Zhenjiang Jiangsu 212013 , China)

机构地区：[1]江苏大学医学院检验医学研究所,江苏镇江212013

出　　处：《江苏大学学报（医学版）》2016年第4期324-327,共4页Journal of Jiangsu University：Medicine Edition

基　　金：国家自然科学基金资助项目(31470881);教育部高等学校博士学科点专项科研基金资助项目(20133227110008);江苏省卫生厅医学科研项目(Z201313)

摘　　要：目的:制备腺病毒AdE-GFP-hTSHR289,并进行鉴定。方法:限制性内切酶NotⅠ和XhoⅠ双酶切分别处理p BluescriptⅡSK(+)-h TSHR289和pAdTrack-CMV,回收纯化目的片段hTSHR289,将其插入至线性化的pAdTrackCMV大片段中;限制性内切酶PmeⅠ线性化pAdTrack-hTSHR289与大肠埃希菌BJ5183中pAdEasy-1质粒同源重组,限制性内切酶PacⅠ酶切验证;将阳性重组质粒转染至人胚肾293A细胞,以包装获得表达hTSHR289的腺病毒AdEGFP-hTSHR289;TCID50法检测重组腺病毒滴度。结果:成功地将hTSHR289重组至腺病毒载体,获得携带hTSHR289重组腺病毒载体,包装出携带hTSHR289基因的腺病毒,滴度达3.5×109pfu/m L。结论:制备得到携带h TSHR289的重组腺病毒。Objective: To develop and identify adenovirus AdE-GFP-hTSHR289. Methods: The hT-5ffi?289 gene was obtained by double digestion pBluescript II SK( + ) -hTSHR289 and pAdTrack- CMV with Not I /Xho I , and inserted into predigested plasmid pAdTrack-CMV. The linearized plasmid pAdTrack-hTSHR289 was transfected into E. coli BJ5183 containing pAdEasy-1. The recombinant plasmidpAdEasy-GFP-hTSHR289 was transfected into 293A cells for getting recombinant adenovirus AdE-GFP-hTSHR289.The virus titer was detected by TCID50 method. Results: hTSHR289 was successfully inserted into adenovirus vector, and obtained the recombinant adenovirus vector. The adenovirus carrying hTSHR2S9 gene were acquired by packaging, the titer of adenovirus was about 3.5 X 109 pfu/ml. Conclusion; The recombinantadenovirus carrying hTSHR2S9 were sucessfully developed.

关 键 词：促甲状腺激素受体 腺病毒 病毒滴度 

分 类 号：R392.1[医药卫生—免疫学]