哈萨克绵羊MC1R和ASIP基因多态性及表达量与被毛颜色表型相关性的研究

摘要/Abstract

摘要：

旨在探究黑色素皮质受体-1基因（MC1R）和刺鼠基因（ASIP）多态性及其mRNA的表达量与哈萨克绵羊被毛颜色之间的关系。本研究利用PCR-SSCP和DNA测序技术，对168只哈萨克绵羊（62只黑色被毛，56只白色被毛及50只棕色被毛）的MC1R和ASIP基因的多态性进行检测，并分析突变位点与不同毛色之间的相关性，同时利用实时定量PCR技术检测3种毛色皮肤组织中这2个基因的表达量，分析基因的表达量与不同毛色之间的相关性。结果显示，MC1R基因的多态位点（P.M73K）与被毛颜色极显著的相关（P<0.01）；ASIP基因的多态位点与被毛颜色没有相关性（P>0.05）。实时定量PCR检测黑色被毛中MC1R基因的表达量与白色被毛中差异极显著（P<0.01），与棕色被毛差异显著（P<0.05），ASIP基因的表达量在不同颜色被毛中没有显著差异（P>0.05）。该研究初步表明，MC1R基因是影响哈萨克绵羊被毛颜色的主效基因。

关键词: 哈萨克绵羊, MC1R, ASIP, 被毛颜色, PCR-SSCP, 实时定量PCR

Abstract:

To investigate the relationship between polymorphism and mRNA expression levels of the two genes (MC1R and ASIP) and coat color in Kazakh sheep, the polymorphism of MC1R and ASIP genes were detected in 168 Kazakh sheep (62 black, 56 white and 50 brown) by PCR-SSCP and sequencing technology, and the relationship between the mutation sites of the two genes and different coat colors were analyzed in sheep. Simultaneously, the real-time PCR was used to detect the expression levels of the two genes in three coat color of skin tissue，their association with coat color was analyzed. The results showed that there was a significant correlation between the MC1R gene polymorphism locus (P.M73K) and coat color (P<0.01), whereas, ASIP gene polymorphic loci was not associated with the coat color (P>0.05). Additionally, the expression level of MC1R in the black hair was significantly different with that in white hair (P<0.01) and brown hair (P<0.05), while the ASIP gene expression was not associated with different hair (P>0.05). Results of the present study indicate that MC1R gene is the major gene affecting the Kazakh sheep coat color.

Key words: Kazakh sheep, MC1R, ASIP, coat color, PCR-SSCP, real-time quantitative PCR

中图分类号:

S826
S813.3

李洪涛，曾献存，张文祥，赵海璇，惠文巧，刘钢，罗燕，班谦，赵宗胜，贾斌. 哈萨克绵羊MC1R和ASIP基因多态性及表达量与被毛颜色表型相关性的研究[J]. 畜牧兽医学报.

LI Hong-tao, ZENG Xian-cun, ZHANG Wen-xiang, ZHAO Hai-xuan, HUI Wen-qiao,LIU Gang, LUO Yan, BAN Qian, ZHAO Zong-sheng, JIA Bin. Analysis of the Relationship between Polymorphism/mRNA Expression Levels of MC1R and ASIP and Coat Color Phenotype in Kazakh Sheep[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA.

参考文献

［1］ROBINS L S, NADEAU J H, JOHNSON K R, et al. Pigmentation phenotypes of variant extension locus alleles result from point mutations that alter MSH receptor function ［J］. Cell, 1993, 72: 827-834.

［2］SLOMINSKI A, TOBIN D J, SHIBAHARA S, et al. Melanin pigmentation in mammalian skin and its hormonal regulation ［J］. Physiol Rev, 2004, 84: 1155-1228.

［3］HALL A M, ORLOW S J. Degradation of tyrosinase induced byphenylthiourea occurs following Golgi maturation ［J］. Pigment Cell Res, 2005, 18: 122-129.

［4］OLLMANN M M, LAMOREUX M L, WILSON B D, et al. Interaction of Agouti protein with the melanocortin 1 receptor in vitro and in vivo ［J］. Genes Dev, 1998, 12: 316-330.

［5］FONTANESI L, DALL’OLIO S, BERETTI F, et al. Coat colours in the Massese sheep breed are associated with mutations in the agouti signalling protein (ASIP) and melanocortin 1 receptor (MC1R) genes ［J］. Animal, 2011, 5(1): 8-17.

［6］NORRIS B J, WHAN V A. A gene duplication affecting expression of the ovine ASIP gene is responsible for white and black sheep［J］. Genome Res, 2008, 18: 1282-1293.

［7］ROYO L J, ALVAREZ I, ARRANZ J J, et al. Differences in the expression of the ASIP gene are involved in the recessive black coat colour pattern in sheep: evidence from the rare Xalda sheep breed ［J］. Anim Genet,2008, 39: 290-293.

［8］GRATTEN J, BERALDI D, LOWDER B V, et al. Compelling evidence that a single nucleotide substitution in TYRP1 is responsible for coat-colour polymorphism in a free-living population of Soay sheep ［J］. Proc Royal Soc Bio Sci, 2007, 274: 619-626.

［9］任玉红, 任彬, 范瑞文, 等. 羊驼MC1R基因克隆及其在不同毛色个体中表达水平研究［J］. 畜牧兽医学报, 2009, 40(6):841-845.

［10］VALVERDE P, HEALY E, JACKSON I, et al. Variants of the melanocyte-stimulating hormone receptor gene are associated with red hair and fair skin in humans ［J］. Nat Genet, 1995, 11: 328-330.

［11］KLUNGLAND H, VAGE D I, GOMEZ-RAYA L, et al. The role of melanocyte-stimulating hormone (MSH) receptor in bovine coat color determination ［J］. Mamm Genome, 1995, 6: 636-639.

［12］MARKLUND L, JOHANSSON MOLLER M, SANDBERG K, et al. Amissense mutation in the gene for melanocyte-stimulating hormone receptor (MC1R) is associated with the chestnut coat color in horses ［J］. Mamm Genome, 1996, 7: 895-899.

［13］TAKEUCHI S, SUZUKI H, YABUUCHI M, et al. A possible involvement of melanocortin 1-receptor in regulating feather color pigmentation in the chicken ［J］. Biochim Biophys Acta, 1996, 1308: 164-168.

［14］VAGE D I, LU D S, KLUNGLAND H, et al. A non-epistatic interaction of agouti and extension in the fox, Vulpes vulpes ［J］. Nat Genet, 1997, 15: 311-315.

［15］KIJAS J M H, WALES R, TORNSTEN A, et al. Melanocortin receptor 1 (MC1R) mutations and coat color in pigs ［J］. Genetics, 1998, 150: 1177-1185.

［16］VAGE D I, KLUNGLAND H, LU D, et al. Molecular and pharmacological characterization of dominant black coat color in sheep ［J］. Mamm Genome, 1999, 10: 39-43.

［17］NEWTON J M, WILKIE A L, HE L, et al. Melanocortin 1 receptor variation in the domestic dog ［J］. Mamm Genome, 2000, 11: 24-30.

［18］MUNDY N I, BADCOCK N S, HART T,et al. Conserved genetic basis of a quantitative plumage trait involved in mate choice ［J］. Science, 2004, 303: 1870-1873.

［19］FONTANESI L, BERETTI F, RIGGIO V, et al. Missense and nonsense mutations in melanocortin 1 receptor (MC1R) gene of different goat breeds: association with red and black coat colour phenotypes but with unexpected evidences ［J］. BMC Genet, 2009, 10: 47.

［20］NACHMAN M W, HOEKSTRA H E, D’AGOSTINO S L, et al. The genetic basis of adaptive melanism in pocket mice ［J］. Proc Natl Acad Sci USA, 2003, 100: 5268-5273.

［21］KERNS J A, OLIVIER M, LUST G, et al. Exclusion of melanocortin-1 receptor (Mc1r) and agouti as candidates for dominant black in dogs ［J］. J Hered, 2003, 94: 75-79.

［22］MUNDY N I, KELLY J. Evolution of a pigmentation gene, the melanocortin-1 receptor in primates ［J］. Am J Phys Anthropol, 2003, 121: 67-80.

［23］BULTMAN S J, MICHAUD E J, WOYCHIK R P. Molecular characterization of the mouse agouti locus ［J］. Cell, 1992, 71: 1195-1204.

［24］RIEDER S, TAOURIT S, MARIAT D, et al. Mutations in the agouti (ASIP), the extension (MC1R), and the brown (TYRP1) loci and their association to coat color phenotypes in horses (Equus caballus) ［J］. Mamm Genome, 2001, 12: 450-455.

［25］KERNS J A, NEWTON J, BERRYERE T G, et al. Characterization of the dog Agouti gene and a nonagouti mutation in German Shepherd dogs ［J］. Mamm Genome, 2004, 15: 798-808.

［26］DROGEMULLER C, GIESE A, MARTINS-WESS F, et al. The mutation causing the black-and-tan pigmentation phenotype of Mangalitza pigs maps to the porcine ASIP locus but does not affect its coding sequence ［J］. Mamm Genome, 2006, 17: 58-66.

［27］GIRARDOT M, GUIBERT S, LAFORET M P, et al. The insertion of a full-length bos taurus LINE element is responsible for a transcriptional deregulation of the Normande Agouti gene ［J］. Pigment Cell Res, 2006, 19: 346-355.

［28］SMIT M A, SHAY T L, BEEVER J E, et al. Identification of an agouti-like locus in sheep ［J］. Anim Genet, 2002, 33: 383-385.

［29］GRATTEN J, PILKINGTON J G, BROWN E A, et al. The genetic basis of recessive self-colour pattern in a wild sheep population ［J］. Heredity, 2010, 104: 206-214.

［30］KERNS J A, CARGILL E J, CLARK L A, et al. Linkage and segregation analysis of black and brindle coat color in domestic dogs ［J］. Genetics, 2007, 176: 1679-1689.

［31］RENIERI C, VALBONESI A, LA M V, et al. Inheritance of coat colour in Merino sheep ［J］. Small Rumin Res, 2008, 74: 23-29.

［32］HE X, LI H T, ZHOU Z Y, et al. Production of brown/yellow patches in the SLC7A11 transgenic sheep via testicular injection of transgene ［J］. J Genet Genomics, 2012, 39(6): 281-285.

［33］ABDEL-MALEK Z A, SCOTT M C, FURUMURA M, et al. The melanocortin 1 receptor is the principal mediator of the effects of agouti signaling protein on mammalian melanocytes ［J］. J Cell Sci 2001, 114: 1019-1024.

[1]	赵珊珊, 肖宁, 李志鹏, 潘庆杰, 李桢, 石德顺, 刘庆友. 马MC1R基因多态位点与毛色相关性分析[J]. 畜牧兽医学报, 2018, 49(8): 1605-1616.
[2]	赵珊珊, 肖宁, 李志鹏, 潘庆杰, 李桢, 石德顺, 刘庆友. 马MC1R基因多态位点与毛色相关性分析[J]. 畜牧兽医学报, 2018, 49(8): 1605-1616.
[3]	罗尚星, 范京惠, 刘宝京, 师乾凯, 侯林杉, 左玉柱. 猪丁型冠状病毒与猪流行性腹泻病毒双重实时荧光定量RT-PCR方法的建立和初步应用[J]. 畜牧兽医学报, 2018, 49(4): 852-858.
[4]	罗尚星, 范京惠, 刘宝京, 师乾凯, 侯林杉, 左玉柱. 猪丁型冠状病毒与猪流行性腹泻病毒双重实时荧光定量RT-PCR方法的建立和初步应用[J]. 畜牧兽医学报, 2018, 49(4): 852-858.
[5]	金元昌，陈壮，李玉峰，袁志栋. 抗性选育协同疫苗免疫对鸡羽囊马立克病毒载量的影响[J]. 畜牧兽医学报, 2015, 46(5): 836-840.
[6]	金元昌，陈壮，李玉峰，袁志栋. 抗性选育协同疫苗免疫对鸡羽囊马立克病毒载量的影响[J]. 畜牧兽医学报, 2015, 46(5): 836-840.
[7]	米焱，张伶俐，李海军，杜晨光，曹贵方，王秀梅. Ghrelin参与FSH对牛卵丘细胞体外增殖的调节[J]. 畜牧兽医学报, 2013, 44(8): 1236-1243.
[8]	米焱，张伶俐，李海军，杜晨光，曹贵方，王秀梅. Ghrelin参与FSH对牛卵丘细胞体外增殖的调节[J]. 畜牧兽医学报, 2013, 44(8): 1236-1243.
[9]	高荣琨，陈伟，曲海娥，李建平，张巧灵. 黑素皮质素受体1（MC1R）基因与VC-獭兔毛色关系的研究[J]. 畜牧兽医学报, 2013, 44(6): 888-893.
[10]	高荣琨，陈伟，曲海娥，李建平，张巧灵. 黑素皮质素受体1（MC1R）基因与VC-獭兔毛色关系的研究[J]. 畜牧兽医学报, 2013, 44(6): 888-893.
[11]	高建斌，昝林森，杨宁，李耀坤，皇甫一凡，郝瑞杰，马向辉，付常振，姜碧杰，成功. 秦川牛DKK1基因SNPs检测及其与体尺、肉质性状的关联分析[J]. 畜牧兽医学报, 2013, 44(3): 376-386.
[12]	李洪涛，曾献存，张文祥，赵海璇，惠文巧，刘钢，罗燕，班谦，赵宗胜，贾斌. 哈萨克绵羊MC1R和ASIP基因多态性及表达量与被毛颜色表型相关性的研究[J]. 畜牧兽医学报, 2013, 44(3): 366-375.
[13]	高建斌，昝林森，杨宁，李耀坤，皇甫一凡，郝瑞杰，马向辉，付常振，姜碧杰，成功. 秦川牛DKK1基因SNPs检测及其与体尺、肉质性状的关联分析[J]. 畜牧兽医学报, 2013, 44(3): 376-386.
[14]	郭云雁，程笃学，张龙超，颜华，赵克斌，王立贤，刘文忠，郭慧慧，陈来华. MyoG和c-fos基因多态性及其合并基因型与猪肌纤维性状的关联分析[J]. 畜牧兽医学报, 2013, 44(2): 188-196.
[15]	郭云雁，程笃学，张龙超，颜华，赵克斌，王立贤，刘文忠，郭慧慧，陈来华. MyoG和c-fos基因多态性及其合并基因型与猪肌纤维性状的关联分析[J]. 畜牧兽医学报, 2013, 44(2): 188-196.