Abstract：Signal transducer and activator of transcription 5a (STAT5a), considered as an important candidate gene for milk production traits, plays key roles in the process of development of regulating animal mammary gland. In the present study, seven tissues included heart, liver, spleen, lung, kidney, cerebellum and mammary gland of Tibetan and Small tail han sheep were detected by qRT-PCR to research the tissues distribution and the differences in expression between different sheep breeds and tissues. The P1 and P2 primer amplification regions of STAT5a in three sheep breeds (Tibetan sheep, Hu sheep and Duolang sheep) using PCR-SSCP and sequencing methods to analysis nucleotide sequence variation and population genetic characteristics. The results indicated that STAT5a gene was expressed in all seven tissues and showed significant differences in species and tissues. The expression of mammary gland was significantly higher than other six tissues in Tibetan and Small tail han sheep (P＜0.01). Between these two breeds, the expression of mammary gland in small han sheep was highly than Tibetan sheep(P＜0.05), but the expression of kidney, cerebellum, and liver were lower than Tibetan sheep (P＜0.05). Two SNPs (c.2150-154C＞T and c.2150-50T＞C) and corresponding alleles A and B were detected in P1 primer amplification regions of STAT5a in sheep. The allele B and genotype BB were the dominant allele and genotype and Tibetan population was moderately polymorphic (0.25＜PIC＜0.5), but other populations were low polymorphic (PIC＜0.25). The χ2 values of the three sheep populations were non-significant (P＞0.05), so those populations lay in the Hardy-Weinberg equilibrium. However, there was no polymorphism in the P2 primer amplification region. In conclusion, these results enrich the contents of the sheep genome and provide the basic information for the molecular mechanism of the formation of mammary gland.
[1]鲍斌，房兴堂，陈宏，等．2008．中国荷斯坦牛STAT5A基因多态性与泌乳性状的相关分析[J]．中国农业科学，41（6）：1872-1878．(Bao B, Fang X T, Chen H, et al. 2008. Polymorphisms of STAT5A Gene and Its Association with Milk Performance Traits in Chinese Holstein Cattle [J]. Scientia Agricultura Sinica, 41(6): 1872-1878.)[2]Boetenin D, White R L, Skolnick M, et al. 1980. Construction of a genetic linkage map in man using restriction fragment length polymorphisms [J]. American Journal of Human Genetics, 32(3): 314-331.[3]Brym P, Kaminski S. 2004. New SSCP polymorphism with in bovine STAT5A gene and its associations with milk performance traits [J]. Journal of Applied Genetics, 45(4): 445-452.[4]Buitenhuis M, Coffer P J, Koenderman L. 2004. Signal transducer and activator of transcription 5(STAT5) [J]. Cell Biology. 36(11): 2120-2124.[5]Byun S, Fang Q, Zhou H, et al. 2009. An effective method for silver-staining DNA in large number of polyacrylamide gels [J]. Analytical Biochemistry, 385(1): 174-175.[6]陈杰．2012．家畜生理学[M]．中国农业出版社，北京．pp．55-98．(Chen J. 2012. Animal Physiology [M]. China Agricultural Publishing House, Beijing. pp. 55-98.)[7]Chorev M, Carmel L. 2012. The function of introns [J]. Frontiers in Genetics, 3(55): 1-15.[8]Gaunitz F, Heise K, Gebhardt R. 2004. A silencer element in the first intron of the glutamine synthetase gene represses induction by glucocorticoids [J]. Molecular Endocrinology, 18(1): 63-69.[9]Geenty, KG. 1979. Effects of weaning age on export lamb production [J]. Proceedings of the New Zealand Society of Animal Production, 39: 202-210.[10]Gong Hua, Zhou Huitong, Hickford J G. 2011. Diversity of the glycine/tyrosine-rich keratin-associated protein 6 gene(KAP6)family in sheep [J]. Molecular Biology Reports, 38(1): 31 -35.[11]Greenwood TA, Kelsoe JR. 2003. Promoter and intronic variants affect the transcriptional regulation of the human dopamine transporter gene [J]. Genomics, 82(5): 511-520.[12]何峰，孙东晓，余英，等．2007．荷斯坦奶牛STAT5A基因的SNPs检测及其与产奶性状关联分析[J]．畜牧兽医学报，38（4）：326-331．(He F, Sun D X, Yu Y, et al, 2007, SNPs Detection of STAT5A Gene and Association with Milk Production Traits in Holstein Cattle [J]. Acta Veterinaria et Zootechnica Sinica, 38(4): 326-331.)[13]Hickford JG, Forrest RH, Zhou H, et al. 2010. Polymorphisms in the ovine myostatin gene(MSTN)and their association with growth and carcass traits in New Zealand Romney sheep [J]. Animal Genetics, 41(1): 64-72.[14]Hui Li, Huiling Zheng. 2014. Signal transducer and activator of transcription 5a inhibited by pimozide may regulate survival of goat mammary gland epithelial cells by regulating parathyroid hormone-related protein [J]. Gene, 51(2): 279-289.[15]James E, Darnell Jr, Lan M. 1994. Jak-STAT Pathways and Trnascriptional activation in response to IFNs and other extracellular signaling proteins [J]. Science, 264(5164): 1415-1421.[16]蒋克平，薛科邦，李春旺．1985.滩羊泌乳性能测定[J].甘肃畜牧兽医，1（1）：1-2.(Jiang K P, Xue K B, Li C W. 1985 . Test of lactation performance of beach sheep [J]. Journal of Gansu Animal Husbandry and Veterinary Medicine, 1(1): 1-2.)[17]季敏，刘学洪，余长林，等．2013．槟榔江水牛STAT5A基因多态性及其与产奶性状的关联性研究[J]．中国牛业科学，39（4）：29-35．(Ji M, Liu X H, Yu C L, et al. 2013. Association of STAT5a Polymorphism and Milk Production Traits in Binlangjiang Buffalo [J]. China Cattle Science, 39(4): 29-35.) [18]Kenneth J, Thomas D, Schmittgen, et al. 2001. Analysis of Relative Gene Expression Data Using Real-Time Quantitative PCR and the 2-??Ct T Method [J]. Methods, 25(4): 402-408.[19]刘芳．2012．藏羚羊STAT3、HIF-1α和HIF-2α基因表达的生物学意义[D]．博士学位论文，青海大学，导师：格日力．pp．52-68．(Liu F. 2012. The biological significance of STAT3, HIF-1α and HIF-2α gene expression in Tibetan antelope [D]. Doctoral thesis, Qinghai University, tutor: lattice force. pp. 52-68.)[20]Levy D E. 1999. Physiological significance of STAT proteins: investigations through gene disruption in vivo [J]. Cellular and Molecular Life Sciences, 55(1): 1559-1567. [21]刘园峰，王桂芝，李秋梅，等．2013．崂山奶山羊LEP和STAT5a基因对泌乳和生长性状的影响[J]．中国农业科学，46（18）：3946-3954．( Liu Y F, Wang G Z, Liu Q M, et al. 2013. The Effect of LEP and STAT5a Genes on Milk and Growth Traits of Laoshan Dairy Goat [J]. Scientia Agricultura Sinica, 46(18): 3946-3954.) [22]Park YW, Juarez M, Ramosc M, et al. 2007. Physico-chemical characteristics of goat and sheep milk [J]. Small Ruminant Research. 68(1-2): 88-143.[23]Selvaggi M, Albarella S, Dario C, et al. 2017. Association of STAT5A Gene Variants with Milk Production Traits in Agerolese Cattle [J]. Biochemical Genetics, 55(2): 158-167.[24]Schmittney M, Doppler W, Ball RK, et al. 1991. Beta-casein gene promoter activity is regulated by the hormone-mediated relief of transcriptional repression and a mammary-gland-specific nuclear factor [J]. Molecular and Cellular Biology, 11(7): 3745-3755.[25]吴贤锋.2015.山羊STAT3和STAT5A基因SNPs、mRNA表达及甲基化对生产性能的影响[D]．硕士学位论文，西北农林科技大学，导师：蓝贤勇．pp．48-67．(Wu X F. 2015. SNPs, mRNA expression and DNA methylation of STAT3 and STAT5A genes and their influences on production traits in goats [J]. Master's degree thesis, Northwest A & F University, Tutor: Lan X Y. pp. 48-67.)[26]杨在宾，杨维仁，张崇宇，等．1997．小尾寒羊和大尾寒羊产乳性能研究[J]．中国养羊，4（1）：41-42．(Yang Z B, Yang W R, Zhang C Y, et al. 1997. Comparative Study on Milk Performance of Small Tail Han Sheep and Big Tail Han Sheep [J]. Chinese sheep, 4(1): 41-42.)[27]张力，潘林阳，杨诗兴，等．1988．哺乳单羔双羔母湖羊的泌乳量及其泌乳曲线的研究[J]．中国畜牧杂志，6（1）：22-23.(Zhang L, Pan L Y, Yang S X, et al. 1988. Study on Lactation Quantity and Lactating Curve of Lamb of Lambs with Two Lambs [J]. Chinese Journal of Animal Science, 6(1): 22-23.)[28]Zhang B, Zhao G, Lan X, et al. 2012. Polymorphism in GHRH gene and its association with growth traits in Chinese native cattle[J]. Research in Veterinary Science, 92(2): 243-246.[29]Zhou H, Hickford J G, Fang Q. 2006. A two-step procedure for extracting genomic DNA from dried blood spots on filter paper for polymerase chain reaction amplification [J]. Analytical Biochemistry, 354(1): 159-161.[30]朱新书，王宏博，包鹏甲，等．2015．藏绵羊哺乳期羔羊早期补饲培育模式研究[J]．安徽农业科学，43（32）：102-103.(Zhu X S, Wang H B, Bao P J, et al. 2015. Study on the Early Supplementary Feeding Model of Tibetan Sheep Lambs in Lactation Period [J]. Journal of Anhui Agri.Sci, 43(32): 102 -103.)
