Selection of reference genes for gene expression studies in porcine skeletal muscle using SYBR green qPCR.
Quantitative PCR (qPCR) is a method for rapid and reliable quantification of mRNA. Internal controls such as reference genes are used to normalize mRNA levels between different samples for an exact comparison of gene transcription level. However, the expression levels of these reference genes may vary between cell types, developmental stages, species and experimental conditions, thus proper normalization strategy is an important precondition for reliable conclusions. In this study, we explored 10 commonly used reference genes in porcine skeletal muscle using SYBR green qPCR. We used both geNorm and NormFinder to analyze the expression stability and found that PPIA, HPRT and eEF-1γ were suitable internal controls for porcine skeletal muscle. However, PPIA, HPRT and SDHA were suitable for skeletal muscle of western pigs while PPIA, eEF-1γ and HPRT for indigenous Chinese pigs. Normalized qPCR data of ROCK2 were compared with microarray data to evaluate our developed set of reference genes.
Feng
Xiaoting
X
Key Laboratory of Swine Genetics and Breeding of Ministry of Agriculture, Breeding and Reproduction of Ministry of Education, Huazhong Agricultural University. No. 1, Lion Hill Street, Wuhan, Hubei 430070, PR China.
Xiong
Yuanzhu
Y
Qian
Hui
H
Lei
Minggang
M
Xu
Dequan
D
Ren
Zhuqing
Z
D000818:Animals; D020869:Gene Expression Profiling / Q000592:standards*; D018482:Muscle, Skeletal / Q000737:chemistry*; D020411:Oligonucleotide Array Sequence Analysis / Q000592:standards*; D009930:Organic Chemicals / Q000737:chemistry*; D016133:Polymerase Chain Reaction; D012015:Reference Standards; D013552:Swine; D054460:rho-Associated Kinases / Q000235:genetics / Q000378:metabolism
D009930:Organic Chemicals; C098798:SYBR Green II; D054460:rho-Associated Kinases