实时荧光定量PCR研究蛋鸡组织基因表达的内参基因筛选  

Screening of Reference Genes for Gene Expression in Layer Hens Tissues Using Real-time Quantitative PCR

作　　者：张蓉[1] 施晓丽[1] 陈朝桂 文光林 赵忠海[1,2] 

机构地区：[1]贵州大学动物科学学院,贵阳550025 [2]高原山地动物遗传育种与繁殖教育部重点实验室,贵阳550025

出　　处：《中国畜牧兽医》2017年第8期2226-2233,共8页China Animal Husbandry & Veterinary Medicine

基　　金：国家自然科学基金(31560642)

摘　　要：为筛选出蛋鸡不同组织中稳定表达的内参基因,试验以120日龄的海兰灰蛋鸡(Gallus domesticus)为试验动物,选取常见内参基因RPS2、β-actin、GAPDH和HMBS作为候选基因,利用实时荧光定量PCR技术Ct值分析法对4个候选内参基因的相对表达进行测定,利用独立评价软件geNorm和NormFinder对蛋鸡的4个候选内参基因在不同组织(心脏、肝脏、肺脏、肾脏、肠、胫骨、胰脏和子宫)中的表达稳定性进行评价。结果显示,Ct值分析和geNorm程序分析均得出相似的结果,即RPS2基因始终占主导地位,稳定性最强,β-actin基因次之;NormFinder软件分析显示,HMBS基因稳定性最好,最佳组合为HMBS和RPS2基因;当分析不同组织中表达恒定的内参基因时,发现不同组织最稳定的内参基因也不完全相同。由此说明,不同的组织器官和不同的分析方法得出的结论虽然不完全一致,但它们却有着一定的潜在共性,发展趋势有一定相似性,即RPS2和β-actin基因相对稳定性更强。To screen the reference genes of stable expression in different tissues of Hy-line layer（Gallus domesticus）,RPS2,β-actin,GAPDHand HMBSgenes were chosen as candidate gene,the software of geNorm,NormFinder and the Ct value were employed to analyze their stability in eight tissues（heart,liver,lung,kidney,duodenum,tibia,pancreas and uterus）of 120 days of age layers using Real-time quantitative PCR technology.The results showed that RPS2 gene was always dominant stability by the analysis of Ct value and geNorm software,theβ-actingene was followed;The NormFinder software showed that the HMBS gene was relative stability,and HMBSand RPS2 genes were the optimal combination;When analyzed the expression of constant reference gene in different tissues,it found that the most stable reference gene in different tissues was not the same.Therefore,it concluded that different tissues had different optimal gene,but they had the common potential,RPS2 and β-actingenes had the relative expression stability comparing the other gene.

关 键 词：蛋鸡 内参基因 稳定性评价 geNorm软件 NormFinder软件 

分 类 号：Q78[生物学—分子生物学]