民猪外周血单核细胞实时荧光定量PCR内参基因的筛选

发布时间：2018-05-09 01:03

  本文选题：民猪 + 外周血单核细胞　； 参考：《中国畜牧兽医》2017年12期

【摘要】：民猪是能够适应东北地区寒冷环境的地方猪种,但目前对其环境适应性相关基因的研究较少,也没有确定适合此研究所需的实时荧光定量PCR的内参基因。因此,本试验通过研究常用的12个内参基因(B2M、ACTB、RPL11、RPL4、YWHAZ、GAPDH、HPRT1、SDHA、HMBS、IDH3B、TUBB2B和TBP1)在不同环境温度下民猪外周血单核细胞中的表达稳定性,旨在确定合适的内参基因进行相关研究。分别在-25、5、10和30℃采集3头民猪耳静脉血分离出单核细胞,利用geNorm、NormFinder、BestKeeper 3种软件分析12个候选内参基因的Ct值,筛选出表达稳定的基因作为内参基因。经geNorm和NormFinder计算获得各候选基因的M值发现,12个候选基因的表达均相对稳定,而BestKeeper的分析则显示ACTB、GAPDH、SDHA、HPRT1、TBP1、YWHAZ基因的SD值均1,可用作本研究条件下的内参基因,而另外6个基因SD值则1,不符合作为内参基因的标准。综合3种分析的结果,在本研究条件下,TBP1基因的稳定性最高,ACTB、GAPDH、SDHA、HPRT1和YWHAZ基因也符合作为内参基因的标准,都可研究在不同环境温度下民猪外周血单核细胞中基因表达时作为内参基因。
[Abstract]:Minzhou pig is a local pig that can adapt to cold environment in Northeast China, but there are few studies on the genes related to its environmental adaptability, and there is no internal reference gene for real-time fluorescent quantitative PCR which is suitable for this study. Therefore, in this experiment, the expression stability of 12 common domestic reference genes (B2MN, ACTBN, RPL11, RPL4, YWHAZN, GAPDH1, HPRT1) in human peripheral blood mononuclear cells (PBMC) was studied in order to determine the appropriate endogenous genes in human peripheral blood mononuclear cells (PBMC) at different ambient temperatures, so as to determine the expression stability of HMBSIDH3BUBB2B and TBP1 in human peripheral blood mononuclear cells (PBMC). Monocytes were isolated from venous blood of three human pig ears at -25 鈩，

本文编号：1863866