民猪外周血单核细胞实时荧光定量PCR内参基因的筛选

doi:10.16431/j.cnki.1671-7236.2017.12.005

Abstract

Abstract:

Min pig is a local pig breed in Northeast China. It has been well-adapted to the local cold weather,but few genes related to its environmental adaptation have been studied. For studies about environmental adaptation of Min pig on molecular level,it is important to have proper reference genes for quantification of gene expression by quantitative Real-time PCR. In this study,12 reference genes (B2M,ACTB,RPL11,RPL4,YWHAZ,GAPDH,HPRT1,SDHA,HMBS,IDH3B,TUBB2B and TBP1) were evaluated for their potential as the reference gene in Min pig peripheral blood mononuclear cells under different temperatures. Blood samples were collected from 3 Min pigs which were under -25,5,10 and 30℃,respectively. Mononuclear cells were separated using density gradient centrifugation. Statistical algorithms including geNorm,Normfinder and BestKeeper were employed to assess the stabilities of these genes. Analysis of geNorm and Normfinder revealed that all these 12 genes were highly stable. However,ACTB,GAPDH,SDHA,HPRT1,TBP1 and YWHAZ genes (SD<1) were found to be more stable than other six genes (SD>1),of which TBP1 was the most stable one using BestKeeper program. To summarize,ACTB,GAPDH,SDHA,HPRT1, TBP1 and YWHAZ genes were suitable to be the reference genes,with TBP1 was the best one.

Key words: Min pig; peripheral blood mononuclear cells; ambient temperature; quantitative Real-time PCR; reference gene

CLC Number:

Q781

MA Hong, LIU Yu, WANG Liang, WANG Wen-tao, WU Sai-hui, HE Xin-miao, LIU Di. Screening of the Reference Genes for the Study of Peripheral Blood Mononuclear Cells by Quantitative Real-time PCR in Min Pigs[J]. , 2017, 44(12): 3426-3433.

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Screening of the Reference Genes for the Study of Peripheral Blood Mononuclear Cells by Quantitative Real-time PCR in Min Pigs

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