可控表达pGH基因转基因小鼠模型的建立与分析  

Establishment and Analysis of Model of Transgenic Mice Sharing Inducible Expression of pGH

作　　者：张建斌[1,2,3] 刘颖 白立景[3,4] 牟玉莲[3] 韩俊文 ZHANG Jian-bin;LIU Ying;BAI Li-jing;MU Yu-lian;HAN Jun-wen(College of Animal Science and Veterinary Medicine, Shanxi Agricultural University, Taigu 030801, China;College of Animal Science and Veterinary Medicine, Tianjin Agricultural University, Tianjin 300384, China;Institute of Animal Science, Chinese Academy of Agricultural Sciences, Beijing 100193, China;Agricultural Genomics Institute at Shenzhen , Chinese Academy of Agricultural Sciences, Shenzhen 518120, China)

机构地区：[1]山西农业大学动物科技学院,太谷030801 [2]天津农学院动物科学与动物医学学院,天津300384 [3]中国农业科学院北京畜牧兽医研究所,北京100193 [4]中国农业科学院深圳农业基因组研究所,深圳518120

出　　处：《畜牧兽医学报》2018年第6期1145-1153,共9页ACTA VETERINARIA ET ZOOTECHNICA SINICA

基　　金：国家转基因动物新品种培育重大专项(2014zx08006005-005);中国农业科学院科技创新工程(ASTIP-IAS05);国家自然科学基金(31501933)

摘　　要：旨在构建整合型可控表达猪生长激素(porcine growth hormone,pGH)基因转基因小鼠模型,通过控制GH的表达时间和表达量来减少GH过早表达产生的副作用,对于培育高饲料转化率大家畜具有重要研究意义。利用本课题组前期成功构建并在PK15细胞系中验证的整合型诱导表达pGH四环素(Tet-on)载体,通过原核显微注射方法制备转基因小鼠。以Southern blot鉴定为阳性的Founder鼠为基础扩繁培育,对转基因后代鼠3~10周龄通过饮水中添加诱导底物强力霉素(DOX)进行诱导试验;利用RT-qPCR技术检测外源基因表达水平;利用放射性免疫检测方法对小鼠血液中猪生长激素水平进行定量检测,同时称重,评估转入基因对小鼠机体表型的影响。结果表明,转基因小鼠诱导组的体重与转基因小鼠非诱导组和同窝非转基因组小鼠相比显著增大(P<0.05),血清中pGH水平显著上升(P<0.05),说明外源GH有效表达并且调节动物机体的生长。结果提示,该模型的建立有助于深入研究GH对机体发育的影响以及机体中影响GH分泌的分子机制和信号通路,促进GH在家畜生产中的应用,也为制备安全、高效的GH转基因大家畜奠定理论基础。This study aimed to construct an integrated and controllable transgenic mice model with pGH gene expression,which could reduce the side effects caused by premature expression of GH through controlling the time and quantity of GH gene expression,it would become a meaningful research to improve feed conversion of livestock.The transgenic mice were successfully obtained by prokaryotic microinjection method using a vector of Tet-on regulatory expression system,which was constructed successfully and verified through transfected PK15 cell line.Based on Southern blot results,positive Founders were bred for getting offspring.When we got the transgenic offspring,the induction experiment was carried out with the induction substrate doxycycline（DOX）supplementation in the drinking water of the transgenic mice at 3-10 weeks old.RT-qPCR technology was used to detect the level of exogenous genes expression.The radioactive immunoassay method was used to detect porcine growth hormone content in serum of mice.Body weight was weighed to evaluate the effects of pGH gene on phenotype of transgenic mice.The results showed that the body weight of induced transgenic mice was higher than that of noninduced transgenic mice and non transgenic mice in the same litter（P〈0.05）;The level of pGH in serum of transgenic mice was significant higher than that of mice in control（P〈0.05）,which indicated that exogenous GHcould effectively express and regulate the growth of mice body.The establishment of model of pGHtransgenic mice contributed to explore the effect of GH gene on body development and the molecular mechanism and signal pathway of growth hormone secretion.It would promote the application of GHin livestock production;it also provided a theoretical foundation and technical support for the preparation of safe and efficient transgenic livestock with GH gene.

关 键 词：四环素可控表达系统 pGH基因 转基因小鼠 

分 类 号：S814.8[农业科学—畜牧学]