实时荧光定量PCR技术在转基因猪生长激素基因差异表达中的应用  

Application of Real-time Fluorescence Quantitative PCR Technology for Porcine Growth Hormone Gene Differential Expression of Transgenic Pigs

作　　者：刘剑飞[1] 吴明明[1] 蒋亚军[1] 迟晓瑶[1] 曹月胜[1] 张廷荣[1] 孙金海[1] 

机构地区：[1]青岛农业大学科技学院,青岛266109

出　　处：《农业生物技术学报》2012年第8期928-935,共8页Journal of Agricultural Biotechnology

基　　金：国家转基因生物新品种培育重大专项(No.2011ZX08006-003;No.2009ZX08010-006B);国家自然科学基金项目(No.30871778);山东省现代农业产业技术体系生猪创新团队建设项目;山东省"泰山学者"建设工程专项

摘　　要：为确定猪生长激素基因(pGH)在F1代转基因猪体内的遗传稳定性及其在育肥初期表达水平的差异,利用SYBRGreenⅠReal-time PCR法检测精子介导纳米基因载体法获得的F0代母猪与非转基因公猪配种所产的7头转pGH基因猪和9头非转基因猪3月龄生长激素的表达水平。根据16个血液样品荧光定量结果绘制溶解曲线和扩增曲线图谱,并用F=(1+E)﹣ΔΔCt,公式进行Ct均值的分析,得出F1代转pGH基因猪生长激素平均表达水平是同期非转基因猪的1.77倍。结果证实,1)外源pGH基因在转基因猪体内能稳定传代;2)并非每头转基因猪体内pGH基因的表达量均高于非转基因猪;3)在F1代转基因猪体内,pGH的平均表达水平显著高于同期非转基因猪。本实验建立了能够稳定测评家猪生长激素相对基因表达水平的方法,为研究转基因与非转基因动物之间生理指标与定量表达的差异性等提供了科学依据。In order to confirm genetic stability and early fattening expression level difference of foreign porcine growth hormone gene （pGH） in Fl generation of transgenic pigs, growth hormone in 7 trans-pGH gene pigs and 9 non-transgenic pigs of 3-month-old were detected by the advantage of SYBR Green I Real-time PCR method, which were produced by F0 transgenic sows made by nano-gene carrier method induced by sperm and normal boar. According to amplification curve and melting curve maps, Ct mean value was gained through the equation F=（I＋E）. The results showed that pGH average expression level in F1 generation of transgenic pigs was 1.77 times as non-transgenic pigs. Suggesting that： 1） foreign gene pGH could stably passage in transgenic pigs, 2） not every pGH gene＇s expression level in transgenic pig was higher than that of non-transgenic pig, 3） average expression level ofpGH gene in F1 generation transgenic pigs was observably higher than that of non-transgenic pigs in the same period. A set of molecule biology method of evaluating pGH relative expression level stably was built, and provided scientific method for discussing physical signs and quantitative expression between transgenic and non-transgenic animals.

关 键 词：转基因 猪生长激素基因(pGH) β-肌动蛋白基因 实时荧光定量PCR 

分 类 号：S828.2[农业科学—畜牧学]