山羊不同组织及不同发育时期骨骼肌内参基因的表达稳定性分析

doi:10.11843/j.issn.0366-6964.2014.08.004

畜牧兽医学报 ›› 2014, Vol. 45 ›› Issue (8): 1228-1236.doi: 10.11843/j.issn.0366-6964.2014.08.004

山羊不同组织及不同发育时期骨骼肌内参基因的表达稳定性分析

陈利，赵薇，占思远，李丹，李利，仲涛，王林杰，张红平^*

(四川农业大学动物科技学院动物遗传育种研究所，雅安 625014)

收稿日期:2014-01-27 出版日期:2014-08-23 发布日期:2014-08-23
通讯作者: 张红平，教授， E-mail：zhp@sicau.edu.cn
作者简介:陈利（1988-），女，贵州盘县人，硕士生，主要从事动物遗传育种与繁殖研究，E-mail：chenli-anjing@163.com
基金资助:
四川省科技支撑计划(2011NZ0099-36)；国家星火计划重点项目(2012GA810001)

The Expression Stability Analysis of Reference Genes in the Different Tissues and Skeletal Muscle of Different Development Periods in Goat

CHEN Li，ZHAO Wei，ZHAN Si-yuan，LI Dan，LI Li，ZHONG Tao，WANG Lin-jie，ZHANG Hong-ping^*

(Institute of Animal Genetics and Breeding，College of Animal Science and Technology，Sichuan Agricultural University，Ya’an 625014，China)

Received:2014-01-27 Online:2014-08-23 Published:2014-08-23

摘要/Abstract

摘要：

为筛选出山羊不同组织、不同时期骨骼肌及骨骼肌卫星细胞(Skeletal muscle satellite cells，SMSCs)中稳定表达的内参基因，以出生后3日龄的山羊不同组织（心、肝、脾、肺、肾、大脑、大肠、小肠、背最长肌、臂三头肌和半膜肌）和不同时期（3、30、60、90和120 d）的3种骨骼肌（背最长肌、臂三头肌和半膜肌）以及培养至不同阶段(1、3、4和7 d) 的山羊SMSCs为研究对象，运用qRT-PCR 技术及geNorm分析软件，探索9个内参基因（ACTIN、GAPDH、TOP2β、YWHAZ、SDHA、PGK1、RP2、RPL4和HPRT1）mRNA的表达稳定性。结果表明，在同一时期的11个组织中，内参基因表达稳定度的排序：HPRT1>RP2>TOP2β>SDHA>GAPDH>ACTIN>YWHAZ>PGK1>RPL4，需要2个内参基因 (HPRT1和RP2) 共同校正目的基因的表达；在不同发育时期的骨骼肌中，各内参的稳定度：YWHAZ>ACTIN>HPRT1>RPL4 >TOP2β>SDHA>PGK1>GAPDH>RP2，需引入3个内参(YWHAZ、ACTIN和HPRT1)才能准确地校正定量结果；在SMSCs中各内参表达的稳定度排序：PGK1>SDHA>ACTIN>RPL4>GAPDH>TOP2β>YWHAZ>RP2>HPRT1，需要3个合适的内参(PGK1、SDHA和ACTIN)作为标准化指标校正定量数据。本试验分别筛选出了能在山羊同一时期不同组织、不同发育时期的骨骼肌及SMSCs中稳定表达的内参基因及需要引入的内参数目，有利于更加准确地校正基因mRNA水平的表达量，为更好地研究基因功能奠定了基础。

Abstract:

To screen the reference genes of stable expression in different tissues，skeletal muscle of different development periods and skeletal muscle satellite cells (SMSCs) in goat，the expression levels of 9 reference genes of ACTIN，GAPDH，TOP2β，YWHAZ，SDHA，PGK1，RP2，RPL4 and HPRT1 mRNA were detected in different tissues (heart，liver，spleen，lung，kidney，brain，small intestine，large intestine,longissimus dorsi，triceps brachii and semimembranosus ) of 3-day-old postnatal kids，and the longissimus dorsi，triceps brachii and semimembranosus of 3，30，60，90 and 120 days old goat and the SMSCs cultured for 1，3，4 and 7 d.The data of expression levels were analyzed using the geNorm software.The results showed that the expressive stability of nine reference genes from high to low were HPRT1，RP2，TOP2β，SDHA，GAPDH，ACTIN，YWHAZ，PGK1，RPL4，and the 2 genes (HPRT1 and RP2) were needed to normalize the qRT-PCR data in different tissues of neonatal goat kids.Meanwhile，the stability of 9 references genes decreased as following: YWHAZ，ACTIN，HPRT1，RPL4，TOP2β，SDHA，PGK1，GAPDH，RP2，and the first 3 genes (YWHAZ，ACTIN and HPRT1) were suggested to accurately measure the true expression levels of target genes during the skeletal muscle of different development stages.Additionally，their expressive stability rank were PGK1，SDHA，ACTIN，RPL4，GAPDH，TOP2β，YWHAZ，RP2，HPRT1 in SMSCs，and 3 genes (PGK1，SDHA and ACTIN) were recommend as standardized factors.Here，the expressive stability of 9 reference genes and suitable genes were confirmed in different tissues，the skeletal muscle of different development periods and the SMSCs in goat.This will be beneficial to the research about the expression and function of gene at mRNA level.

中图分类号:

S827
Q786

陈利，赵薇，占思远，李丹，李利，仲涛，王林杰，张红平. 山羊不同组织及不同发育时期骨骼肌内参基因的表达稳定性分析[J]. 畜牧兽医学报, 2014, 45(8): 1228-1236.

CHEN Li，ZHAO Wei，ZHAN Si-yuan，LI Dan，LI Li，ZHONG Tao，WANG Lin-jie，ZHANG Hong-ping. The Expression Stability Analysis of Reference Genes in the Different Tissues and Skeletal Muscle of Different Development Periods in Goat[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2014, 45(8): 1228-1236.

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山羊不同组织及不同发育时期骨骼肌内参基因的表达稳定性分析

The Expression Stability Analysis of Reference Genes in the Different Tissues and Skeletal Muscle of Different Development Periods in Goat

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