狐狸MC1R基因编码区c.40A>C和c.41C>T相邻变异研究

doi:doi: 10.11843/j.issn.0366-6964.2016.10.010

摘要/Abstract

摘要：

为了检测狐狸MC1R基因多态性及其与毛色表型的相关性，本研究采集两个狐属12种毛色共计163只狐狸的皮肤组织样，利用PCR扩增和产物直接测序的方法获得狐狸MC1R基因1 054 bp长的核苷酸序列，并进行了SNPs筛查。用PopGen32和SHEsis软件对突变位点进行了群体遗传学分析，用PANTHER软件评估了突变对基因产生的功能影响，用SPSS二元变量相关统计方法分析了多态位点与毛色表型间的相关性。结果表明，狐狸MC1R基因编码区40（c.40A>C）和41 位点（c.41C>T）存在2 个相邻错义突变，导致其编码的第14位氨基酸发生了变异：当第40 位点为A时，氨基酸由苏氨酸（Thr）转变为异亮氨酸（Ile）；当第40 位点为C时，氨基酸由脯氨酸（Pro）转变为亮氨酸（Leu）。北极狐属狐在41 位点基因型全部为TT型，而狐属狐大部分个体均为CC型，不存在TT型，推测该位点可能是区分狐狸属间的一个重要功能位点。PANTHER预测获知第41 位点突变导致的氨基酸替换（p.Pro14Leu）对MC1R功能有显著影响。SPSS二元变量相关分析结果表明，41 位点多态性与狐狸毛色表型存在显著低度相关性，推测狐狸MC1R基因编码区第41位点可能是参与其毛色形成的一个相对重要功能位点。

关键词: 狐狸, MC1R基因, 40和41位点, 多态性, 毛色

Abstract:

In order to detect the relationship between polymorphism of MC1R gene and coat color in fox，a total of 163 skin samples of 12 coat color foxes were collected.The nucleotide sequence (1 054 bp) of fox MC1R gene were obtained by the method of PCR and direct sequencing，and the polymorphism were analyzed.The population genetics were analyzed using PopGen32 and SHEsis softwares.The effect of mutations on the function of MC1R gene was evaluated using PANTHER software.The relationships between the variable sites and coat color were analyzed by the statistical methods of SPSS bivarate correlation analysis.Two adjacent missense mutations (c.40A>C and c.41C>T) were found in the coding region of fox MC1R gene，which resulted in codon change of p.Thr14Ile or p.Pro14Leu.When 40 site was A，it led to the substitution between threonine (Thr) and isoleucine (Ile).When 40 site was C，it led to the substitution between proline (Pro) and leucine (Leu).All the genotypes of two coat color foxes belonging to Alopex were TT.However，the genotypes of most coat color foxes belonging to Vulpes were CC.It was supposed that 41 site was important in distinguishing Alopex and Vulpes.The in silico functional analysis showed that the amino acid substitution at p.Pro14Leu had significant impact on the function of MC1R.The statistical analysis showed the polymorphism of 41 site had significant low correlation with the fox coat color.The results indicate that SNP c.41C>T in the coding region of the MC1R gene is probably associated with the coat color in fox. In order to detect the relationship between polymorphism of MC1R gene and coat color in fox，a total of 163 skin samples of 12 coat color foxes were collected.The nucleotide sequence (1 054 bp) of fox MC1R gene were obtained by the method of PCR and direct sequencing，and the polymorphism were analyzed.The population genetics were analyzed using PopGen32 and SHEsis softwares.The effect of mutations on the function of MC1R gene was evaluated using PANTHER software.The relationships between the variable sites and coat color were analyzed by the statistical methods of SPSS bivarate correlation analysis.Two adjacent missense mutations (c.40A>C and c.41C>T) were found in the coding region of fox MC1R gene，which resulted in codon change of p.Thr14Ile or p.Pro14Leu.When 40 site was A，it led to the substitution between threonine (Thr) and isoleucine (Ile).When 40 site was C，it led to the substitution between proline (Pro) and leucine (Leu).All the genotypes of two coat color foxes belonging to Alopex were TT.However，the genotypes of most coat color foxes belonging to Vulpes were CC.It was supposed that 41 site was important in distinguishing Alopex and Vulpes.The in silico functional analysis showed that the amino acid substitution at p.Pro14Leu had significant impact on the function of MC1R.The statistical analysis showed the polymorphism of 41 site had significant low correlation with the fox coat color.The results indicate that SNP c.41C>T in the coding region of the MC1R gene is probably associated with the coat color in fox.

Key words: fox, MC1R gene, 40 and 41 sites, polymorphism, coat color

中图分类号:

徐桂利，张文香，段玲欣，巩元芳，葛慕湘，刘谢荣，王书朋，果新苓，刘铮铸. 狐狸MC1R基因编码区c.40A>C和c.41C>T相邻变异研究[J]. 畜牧兽医学报, 2016, 47(10): 2020-2026.

XU Gui-li，ZHANG Wen-xiang，DUAN Ling-xin，GONG Yuan-fang，GE Mu-xiang，LIU Xie-rong，WANG Shu-peng，GUO Xin-ling，LIU Zheng-zhu. Study on Adjacent Variation of c.40A>C and c.41C>T of Fox MC1R Gene in Different Populations[J]. ACTA VETERINARIA ET ZOOTECHNICA SINICA, 2016, 47(10): 2020-2026.

参考文献

［1］BENNETT D C，LAMOREUX M L.The color loci of mice-a genet iccentury［J］.Pigment Cell Res，2003，16(4)：333-344.
［2］马泽芳，崔凯.貂狐貉实用养殖技术［M］.北京：中国农业出版社，2014.
MA Z F，CUI K.Practical breeding techniques of mink，fox and raccoon dog［M］.Beijing：China Agriculture Press，2014.（in Chinese）
［3］佟煜仁，张志明.毛皮动物毛色遗传及繁育新技术［M］.北京：金盾出版社，2009.
TONG Y R，ZHANG Z M.New technology for coat color genetics and breeding of fur animal［M］.Beijing：Jindun Press，2009.（in Chinese）
［4］VÅGE D I，LU D，KLUNGLAND H，et al.A non-epistatic interaction of agouti and extension in the fox，Vulpes vulpes［J］.Nat Genet，1997，15(3)：311-315.
［5］VÅGE D I，FUGLEI E，SNIPSTAD K，et al.Two cysteine substitutions in the MC1R generate the blue variant of the Arctic fox (Alopex lagopus) and prevent expression of the white winter coat［J］.Peptides，2005，26(10)：1814-1817.
［6］PALMER J S，DUFFY D L，BOX N F，et al.Melanocortin-1 receptor polymorphisms and risk of melanoma：is the association explained solely by pigmentation phenotype?［J］.Am J Hum Genet，2000，66(1)：176-186.
［7］BASHMAKOVA E E，KRASITSKAYA V V，BONDAR A A，et al.Bioluminescent assay to detect melanocortin-1 receptor (MC1R) polymorphisms (R160W，R151C，and D294H)［J］.Mol Biol (Mosk)，2015，49(6)：953-958.
［8］ABITBOL M，LEGRAND R，TIRET L.A missense mutation in melanocortin 1 receptor is associated with the red coat colour in donkeys［J］.Anim Genet，2014，45(6)：878-880.
［9］李洪涛，曾献存，张文祥，等.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.
LI H T，ZENG X C，ZHANG W X，et al.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，2013，44(3)：366-375.（in Chinese）
［10］郭多，巴彩凤，苏玉虹，等.犬MC1R基因R306ter与毛色性状相关性研究［J］.中国实验动物学报，2004，12(2)：100-102.
GUO D，BA C F，SU Y H，et al.Study on the relationship between r306ter loci in mc1r gene and coat color trait in dogs［J］.Acta Laboratorium Animalis Scientia Sinica，2004，12(2)：100-102.（in Chinese）
［11］SAMBROOK J，FRITSCH E F，MANIATIS T.Molecular cloning［M］.New York：Cold Spring Habor Laboratory Press，1989.
［12］HALL T A.BioEdit：A user-friendly biological sequences alignment editor and analysis program for Windows 95/98/NT［J］.Nucleic Acids Symp Ser，1999，41：95-98.
［13］YEH F C，YANG R C，BOYLE T.POPGENE version 1.31：Microsoft Windows-based freeware for population genetic analysis-Quick User Guide［M］.University of Alberta and Centre for International Forestry Research.Edmonton，AB Canada，1999：1-28.
［14］SHI Y Y，HE L.SHEsis，a powerful software platform for analyses of linkage disequilibrium，haplotype construction，and genetic association at polymorphism loci［J］.Cell Res，2005，15(2)：97-98.
［15］BRUNHAM L R，SINGARAJA R R，PAPE T D，et al.Accurate prediction of the functional significance of single nucleotide polymorphisms and mutations in the ABCA1 gene［J］.PLoS Genet，2005，1(6)：e83.
［16］THOMAS P D，KEJARIWAL A，GUO N，et al.Applications for protein sequence-function evolution data：mRNA/protein expression analysis and coding SNP scoring tools［J］.Nucleic Acids Res，34 (Web Server issue)：W645-650.
［17］苗永旺，吴桂生，王磊，等.黑色素皮质素受体1 基因对水牛毛色遗传的影响［J］.中国科学C辑：生命科学，2009，39(12)：1155-1161.
MIAO Y W，WU G S，WANG L，et al.The role of MC1R gene in buffalo coat color［J］.Science in China(Series C：Life Sciences)，2009，39(12)：1155-1161.（in Chinese）
［18］孙逸敏.利用SPSS软件分析变量间的相关性［J］.新疆教育学院学报，2007，23(2)：120-123.
SUN Y M.Using SPSS software to analyze the correlation between variables［J］.Journal of Xinjiang Education Institute，2007，23(2)：120-123.（in Chinese）
［19］NEWTON J M，WILKIE A L，HE L，et al.Melanocortin 1 receptor variation in the domestic dog［J］.Mamm Genome，2000，11(1)：24-30.
［20］WANG G D，CHENG L G，FAN R X，et al.Signature of balancing selection at the MC1R gene in Kunming dog populations［J］.PLoS One，2013，8(2)：e55469.
［21］DREGER D L，SCHMUTZ S M.A new mutation in MC1R explains a coat color phenotype in 2 "old" breeds：Saluki and Afghan hound［J］.J Hered，2010，101(5)：644-649.
［22］NOWACKA-WOSZUK J，SALAMON S，GORNA A，et al.Missense polymorphisms in the MC1R gene of dog，red fox，arctic fox and Chinese raccon dog［J］.J Anim Breed Genet，2013，130(2)：136-141.

[1]	李青, 王慧华, 卢曾奎, 金美林, 储明星, 魏彩虹. 绵羊PDGF-D基因多态性与尾型的关联[J]. 畜牧兽医学报, 2019, 50(4): 688-700.
[2]	李青, 王慧华, 卢曾奎, 金美林, 储明星, 魏彩虹. 绵羊PDGF-D基因多态性与尾型的关联[J]. 畜牧兽医学报, 2019, 50(4): 688-700.
[3]	杜小龙, 王麒, 张乐超, 葛琳涵, 刘小辉, 王涵, 李兰会, 李祥龙. 山羊DCT基因启动子区甲基化水平、SNP与毛色特征关系研究[J]. 畜牧兽医学报, 2019, 50(2): 271-279.
[4]	杜小龙, 王麒, 张乐超, 葛琳涵, 刘小辉, 王涵, 李兰会, 李祥龙. 山羊DCT基因启动子区甲基化水平、SNP与毛色特征关系研究[J]. 畜牧兽医学报, 2019, 50(2): 271-279.
[5]	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.
[6]	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.
[7]	唐朋, 凌笑笑, 高泽成, 贾聪俊, 梁春年, 吴晓云, 褚敏, 阎萍. 不同毛色牦牛皮肤组织学观察及MC1R基因功能验证[J]. 畜牧兽医学报, 2018, 49(7): 1377-1386.
[8]	唐朋, 凌笑笑, 高泽成, 贾聪俊, 梁春年, 吴晓云, 褚敏, 阎萍. 不同毛色牦牛皮肤组织学观察及MC1R基因功能验证[J]. 畜牧兽医学报, 2018, 49(7): 1377-1386.
[9]	王建文, 李林玲, 王镜力, 孟军, 曾亚琦, 姚新奎, 辛雅莉. 伊犁马ACTN3基因外显子多态性研究[J]. 畜牧兽医学报, 2018, 49(6): 1299-1306.
[10]	王建文, 李林玲, 王镜力, 孟军, 曾亚琦, 姚新奎, 辛雅莉. 伊犁马ACTN3基因外显子多态性研究[J]. 畜牧兽医学报, 2018, 49(6): 1299-1306.
[11]	张文建, 马广伟, 张潇飞, 褚衍凯, 刘静, 杨华, 闫晓红, 李玉茂, 李辉, 王宁. 羊毛品质性状相关基因CCNY的验证和分析[J]. 畜牧兽医学报, 2017, 48(8): 1389-1400.
[12]	张文建, 马广伟, 张潇飞, 褚衍凯, 刘静, 杨华, 闫晓红, 李玉茂, 李辉, 王宁. 羊毛品质性状相关基因CCNY的验证和分析[J]. 畜牧兽医学报, 2017, 48(8): 1389-1400.
[13]	海汀, 柴志欣, 钟金城. 麦洼牦牛GH、GHR、GHSR基因的SNPs检测及其与体尺性状的关联分析[J]. 畜牧兽医学报, 2017, 48(4): 605-617.
[14]	海汀, 柴志欣, 钟金城. 麦洼牦牛GH、GHR、GHSR基因的SNPs检测及其与体尺性状的关联分析[J]. 畜牧兽医学报, 2017, 48(4): 605-617.
[15]	孙笑尉，张雪莲，李振，庞全海. 染色质重塑因子BPTF、BRG1在不同毛色绵羊皮肤的表达与定位[J]. 畜牧兽医学报, 2017, 48(3): 446-453.