title: Technical note: Selection of suitable reference genes for studying gene expression in milk somatic cell of yak (Bos grunniens) during the lactation cycle.
abstract: Quantitative real-time PCR is the most sensitive technique for gene expression analysis. Data normalization is essential to correct for potential errors incurred in all steps from RNA isolation to PCR amplification. The commonly accepted approach for normalization is the use of reference gene. Until now, no suitable reference genes have been available for data normalization of gene expression in milk somatic cells of lactating yaks across lactation. In the present study, we evaluated the transcriptional stability of 10 candidate reference genes in milk somatic cells of lactating yak, including ACTB, B2M, GAPDH, GTP, MRPL39, PPP1R11, RPS9, RPS15, UXT, and RN18S1. Four genes, RPS9, PPP1R11, UXT, and MRPL39, were identified as being the most stable genes in milk somatic cells of lactating yak. Using the combination of RPS9, PPP1R11, UXT, and MRPL39 as reference genes, we further assessed the relative expression of 4 genes of interest in milk somatic cells of yak across lactation, including ELF5, ABCG2, SREBF2, and DGAT1. Compared with expression in colostrum, the overall transcription levels of ELF5, ABCG2, and SREBF2 in milk were found to be significantly upregulated in early, peak, and late lactation, and significantly downregulated thereafter, before the dry period. A similar pattern was observed in the relative expression of DGAT1, but no significant difference was revealed in its expression in milk from late lactation compared with colostrum. Based on these results, we suggest that the geometric mean of RPS9, PPP1R11, UXT, and MRPL39 can be used for normalization of real-time PCR data in milk somatic cells of lactating yak, if similar experiments are performed.
authors.affiliation: College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang 110866, China.
authors.forename: W L
authors.initials: WL
authors.lastname: Bai
authors.affiliation: College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang 110866, China. Electronic address: yinronghuan@163.com.
authors.forename: R H
authors.initials: RH
authors.lastname: Yin
authors.affiliation: Institute of Biotechnology, Animal Science Research Academy of Sichuan Province, Chengdu 610066, China.
authors.forename: S J
authors.initials: SJ
authors.lastname: Zhao
authors.affiliation: College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang 110866, China.
authors.forename: W Q
authors.initials: WQ
authors.lastname: Jiang
authors.affiliation: Research Academy of Animal Husbandry and Veterinary Medicine Sciences of Jilin Province, Changchun 130062, China.
authors.forename: R L
authors.initials: RL
authors.lastname: Yin
authors.affiliation: Academy of Animal Science and Veterinary Medicine, Qinghai University, Xining 810016, China.
authors.forename: Z J
authors.initials: ZJ
authors.lastname: Ma
authors.affiliation: College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang 110866, China.
authors.forename: Z Y
authors.initials: ZY
authors.lastname: Wang
authors.affiliation: College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang 110866, China.
authors.forename: Y B
authors.initials: YB
authors.lastname: Zhu
authors.affiliation: College of Animal Science and Veterinary Medicine, Shenyang Agricultural University, Shenyang 110866, China.
authors.forename: G B
authors.initials: GB
authors.lastname: Luo
authors.affiliation: College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, China.
authors.forename: R J
authors.initials: RJ
authors.lastname: Yang
authors.affiliation: College of Animal Science and Veterinary Medicine, Jilin University, Changchun 130062, China. Electronic address: zhzhao@jlu.edu.cn.
authors.forename: Z H
authors.initials: ZH
authors.lastname: Zhao
mesh_terms: D000818:Animals; D002417:Cattle / Q000502:physiology*; D003126:Colostrum; D005260:Female; D005786:Gene Expression Regulation / Q000502:physiology*; D007774:Lactation / Q000235:genetics / Q000502:physiology*; D008892:Milk / Q000166:cytology; D016133:Polymerase Chain Reaction; D011247:Pregnancy; D012313:RNA / Q000235:genetics; D060888:Real-Time Polymerase Chain Reaction
chemical_list: D012313:RNA
keywords: gene expression; milk somatic cell; reference gene; yak