银黑狐MC1R基因核心启动子区的鉴定

doi:10.16431/j.cnki.1671-7236.2022.07.002

Abstract

Abstract: 【Objective】 This study was aimed to screen the core promoter region of melanocortin 1 receptor (MC1R) gene of silver fox,and provide a theoretical basis for exploring the expression regulatory mechanism of MC1R gene.【Method】 Using genomic DNA of silver fox as template,the sequence of the 5'-UTR region of MC1R gene was obtained by PCR amplification.The active region of the MC1R gene promoter of silver fox was predicted comprehensively using 3 online biology softwares.The deletion fragments of different lengths in 5'-UTR region of MC1R gene were amplified by PCR,which were cloned into pMD19-T vector,and the recombinant plasmid was transfected into melanin B16 cells.The luciferase activities of 10 missing fragments were detected by dual luciferase detection technique.【Result】 The 5'-UTR region sequence of MC1R gene in silver fox was successfully obtained,and ―596/+73 bp might be the active region of the promoter using biology softwares prediction.The detection of dual luciferase activity revealed that the constructed luciferase expression vectors of 10 different deletion fragments all had promoter activity.Among them,pGL3-MC1RP8 (―520/+73 bp) had strong activity,indicating that it was the core promoter region,which was basically consistent with the software prediction results.【Conclusion】 The core promoter region of MC1R gene in silver fox was determined to be ―520/+73 bp,which laid a theoretical foundation for the further study on coat color regulation mechanism of MC1R gene.

Key words: silver fox; MC1R gene; promoter; core region

CLC Number:

LIU Huayun, ZHANG Lei, XU Guili, ZHANG Tianhao, XIE Yuchun, DUAN Lingxin, LIU Zhengzhu, GONG Yuanfang. Identification of the Core Promoter Region of MC1R Gene in Silver Fox[J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(7): 2442-2450.

References

[1] LIU Z Z,GONG Y F,FENG M S,et al.Genetic variations of the coding region of the melanocortin receptor 1(MC1R) gene in the fox[J].Veterinary Dermatology,2016,27(3):135-e36.
[2] 李丽莎,彭永东,郑晓宁,等.山羊PMEL基因启动子活性及转录调控元件分析[J].畜牧兽医学报,2017,48(5):826-835. LI L S,PENG Y D,ZHENG X N,et al.Analysis of the promoter activity and transcriptional regulatory elements of goat PMEL gene[J]. Acta Veterinaria et Zootechnica Sinica,2017,48(5):826-835.(in Chinese)
[3] SUZUKI H.Evolutionary and phylogeographic views on MC1R and Asip variation in mammals[J]. Genes Genes & Genetic Systems,2013,88(3):155-164.
[4] BREIT A,BUCH T R,BOEKHOFF I,et al.Alternative G protein coupling and biased agonism:New insights into melanocortin-4 receptor signalling[J].Molecular and Cellular Endocrinology,2011,331(2):232-240.
[5] 徐桂利,刘铮铸,巩元芳,等.毛皮动物毛色调控基因的研究进展[J].黑龙江畜牧兽医,2015,13:52-54. XU G L,LIU Z Z,GONG Y F,et al.Research progress on the controlling genes for the coat colors of fur animals[J].Heilongjiang Animal Science and Veterinary Medicine,2015,13:52-54.(in Chinese)
[6] 刘玲玲,孟军,菲茹扎,等.伊犁马毛色性状的全基因组关联分析[J].中国畜牧杂志,2021,57(6):116-119. LIU L L,MENG J,FEI R Z,et al.Genome-wide association analysis of Yili horse hair color traits[J].Chinese Journal of Animal Science,2021,57(6):116-119.(in Chinese)
[7] 王晓薇,樊毅,马小明,等.滩羊毛色候选基因MC1R和TCF25的mRNA表达量定量分析[J].浙江农业学报,2020,32(8):1351-1356. WANG X W,FAN Y,MA X M,et al.Quantitative analysis of mRNA expression of Tan sheep coat color candidate genes MC1R and TCF25[J].Journal of Zhejiang Agricultural Sciences,2020,32(8):1351-1356.(in Chinese)
[8] 南瑞鹏,陈娟娟,李振,等.羊驼MC1R真核表达载体的构建及其在毛囊干细胞中的表达[J].农业生物技术学报,2017,25(4):607-613. NAN R P,CHEN J J,LI Z,et al.Construction of eukaryotic expression vector of MC1R gene and its expression in hair follicle stem cells of alpaca (Lama pacos)[J].Journal of Agricultural Biotechnology,2017,25(4):607-613.(in Chinese)
[9] ZHANG Y,XU H Y,MU J H,et al.Inactivation of ADAMTS18 by aberrant promoter hypermethylation contribute to lung cancer progression[J].Journal of Cellular Physiology,2019,234(5):6965-6975.
[10] TOJI N,KOSHI K,FURUSAWA T,et al.A cell-based interferon-tau assay with an interferon-stimulated gene 15 promoter[J].Biomedical Research,2018,39(1):13-20.
[11] SAMBROOKJ,FRITSCH E F,MANIATIS T.Molecular Cloning[M].New York:Spring Habor Laboratory Press,1989.
[12] AOKI H,MORO O.Involvement of microphthalmia-associated transcription factor (MITF) in expression of human melanocortin-1 receptor (MC1R)[J].Life Sciences,2002,71(18):2171-2179.
[13] DREGER D L,SCHMUTZ S M.A SINE insertion causes the black-and-tan and saddle tan phenotypes in domestic dogs[J].The Journal of Heredity, 2011,102(S1):S11-S18.
[14] 宋兴超,杨福合,巴恒星,等.赤狐黑色素皮质激素受体-1基因结构特征预测及分子进化分析[J].中国畜牧兽医,2013,40(1):9-13. SONG X C,YANG F H,BA H X,et al.Prediction on structural characteristics and phylogenetic evolution analysis of MC1R gene in red fox (Vulpes vulpes)[J]. China Animal Husbandry & Veterinary Medicine,2013,40(1):9-13.(in Chinese)
[15] 徐桂利,张文香,段玲欣,等.狐狸MC1R基因编码区c.40 A>C和c.41 C>T相邻变异研究[J].畜牧兽医学报,2016,47(10):2020-2026. XU G L,ZHANG W X,DUAN L X,et al.Study on adjacent variation of c.40 A>C and c.41 C>T of fox MC1R gene in different populations[J].Acta Veterinaria et Zootechnica Sinica,2016,47(10):2020-2026.(in Chinese)
[16] GONG Y F,LIU Z Z,ZHANG W X,et al.Genetic polymorphisms in the 5'-flanking region of the melanocortin 1 receptor (MC1R) gene in foxes[J]. Russian Journal of Genetics,2017,53(4):490-497.
[17] 高心怡,王聪,吴婧婷,等.美洲水貂MC1R基因CDS区克隆与序列分析[J].黑龙江畜牧兽医,2019,11:147-150. GAO X Y,WANG C,WU J T,et al.Cloning and sequence analysis of MC1R gene CDS region in Neovison vision[J].Heilongjiang Animal Science and Veterinary Medicine,2019,11:147-150.(in Chinese)
[18] 刘满清,刘清神,方梅霞,等.沙皮犬MC1R基因的克隆与序列分析[J].广东农业科学,2014,24:127-130. LIU M Q,LIU Q S,FANG M X,et al.Cloning and sequence analysis of MC1R gene in Chinese Shar-pei dog[J].Guangdong Agricultural Sciences,2014,24:127-130.(in Chinese)
[19] 徐超,宋兴超,岳志刚,等.4种犬科动物黑素皮质激素受体1基因序列比对及遗传进化分析[J].中国畜牧兽医,2014,41(1):25-29. XU C,SONG X C,YUE Z G,et al.Sequence comparison and genetic evolution analysis of MC1R gene in four Canidae families[J].China Animal Husbandry & Veterinary Medicine,2014,41(1):25-29.(in Chinese)
[20] 梁爽.4个家兔群体MC1R基因外显子的遗传多样性分析[J].上海农业科技,2016,5:57-59. LIANG S.Analysis of genetic diversity of MC1R gene exons in 4 rabbit populations[J].Shanghai Agricultural Science and Technology,2016,5:57-59.(in Chinese)
[21] 段耀举.家兔毛色候选基因MC1R、AGOUTI、TYR、TYRP1及MLPH的研究分析[D].泰安:山东农业大学,2018. DUAN Y J.Research and analysis of the candidate gene MC1R,AGOUTI,TYR,TYRP1 and MLPH of rabbits' color[D].Tai'an:Shandong Agricultural University,2018.(in Chinese)
[22] 高荣琨,陈伟,曲海娥,等.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(16):888-893. GAO R K,CHEN W,QU H E,et al.Study on the relationship between the melanocortin 1 receptor gene (MC1R) and the coat color inrex-rabbits[J].Acta Veterinaria et Zootechnica Sinica,2013,44(16):888-893.(in Chinese)
[23] 赵珊珊.马MC1R基因多态位点与毛色相关性研究[D].南宁:广西大学,2018. ZHAO S S.Polymorphism loci of MC1R gene and coat color in horses[D].Nanning:Guangxi University,2018.(in Chinese)
[24] 付琳,任航行,王高富,等.酉州乌羊MC1R基因的克隆鉴定、生物信息学及多态性分析[J].核农学报,2021,35(1):31-40. FU L,REN H X,WANG G F,et al.Amplification,bioinformatics and polymorphism analysis of MC1R gene in Youzhou Dark goat[J].Journal of Nuclear Agriculture,2021,35(1):31-40.(in Chinese)
[25] 薛丽娜,罗惠娣,周胜花,等.不同毛色绵羊皮肤组织中MC1R、Agouti及TYR基因差异表达研究[J].中国畜牧杂志,2021,57(1):91-96. XUE L N,LUO H D,ZHOU S H,et al.Expression and regulation analysis of MC1R,Agouti and TYR genes in sheep skins with different coat colors[J].Chinese Journal of Animal Science,2021,57(1):91-96.(in Chinese)
[26] 李隐侠,张俊,钱勇,等.山羊MC1R基因克隆、表达及其多态性研究[J].江苏农业科学,2020,48(1):70-74. LI Y X,ZHANG J,QIAN Y,et al.Cloning,expression and polymorphism of goat MC1R gene[J]. Jiangsu Agricultural Sciences,2020,48(1):70-74.(in Chinese)
[27] 王小女,张晨,岳雨露,等.豫南黑猪MC1R基因序列分析[J].黑龙江畜牧兽医,2021,2:43-48. WANG X N,ZHANG C,YUE Y L,et al.Sequence analysis of MC1R gene in Yunan Black pig[J].Heilongjiang Animal Science and Veterinary Medicine,2021,2:43-48.(in Chinese)
[28] 彭刚.猪MC1R、ASIP、TYR、TYRP1和SLC45A2基因多态性及其与毛色相关性研究[D].武汉:华中农业大学,2020. PENG G.Polymorphisms of MC1R,ASIP,TYR,TYRP1,SLC45A2 gene and their association with coat color in pigs[D].Wuhan:Huazhong Agricultural University,2020.(in Chinese)
[29] JIA Q T,CAO C W,TANG H,et al.A 2 bp insertion (c.67_68insCC) in MC1R causes recessive white coat color in Bama Miniature pigs[J].Journal of Genetics and Genomics,2017,44(4):215-217.
[30] 姚一博.中国地方黄牛3个毛色相关基因(MC1R、TYRP1和ASIP)的多态性研究[D].杨凌:西北农林科技大学,2019. YAO Y B.Polymorphism of three coat colorrelated genes (MC1R,TYRP1 and ASIP) in Chinese local cattle[D].Yangling:Northwest A & F University,2019.(in Chinese)
[31] MIAO Y,WU G,WANG L,et al.The role of MC1R gene in buffalo coat color[J].Science China.Life Sciences,2010,53(2):267-272.
[32] 韩敏振,舒适,黄荣,等.牦牛黑色素相关基因的研究进展[J].青海畜牧兽医杂志,2020,50(5):52-55. HAN M Z,SHU S,HUANG R,et al.Research progress of melanin-related genes in yak[J].Qinghai Journal of Animal Husbandry and Veterinary Medicine,2020,50(5):52-55.(in Chinese)
[33] 林宇纾,蒋恒洁,杨丰硕,等.水牛黑色素皮质素1受体基因克隆、生物信息学分析及表达模式研究[J].中国畜牧兽医,2015,42(10):2529-2537. LIN Y S,JIANG H J,YANG F S,et al.Cloning,bioinformatics and expression pattern analysis of buffalo MC1R gene[J].China Animal Husbandry & Veterinary Medicine,2015,42(10):2529-2537.(in Chinese)
[34] 张秋月,任玉红,范瑞文,等.羊驼MC1R基因的扩增与生物信息学分析[J].山西农业大学学报(自然科学版),2015,35(2):119-123. ZHANG Q Y,REN Y H,FAN R W,et al.Gene amplification and bioinformatics analysis of MC1R in alpaca[J].Journal of Shanxi Agricultural University (Natural Science Edition),2015,35(2):119-123.(in Chinese)
[35] 王婧,李冰,刘翠翠,等.启动子结构和功能研究进展[J].生物技术通报,2014,8:40-45. WANG J,LI B,LIU C C,et al.Advances of the studies on structure and function of promoter[J].Biotechnology Bulletin,2014,8:40-45.(in Chinese)
[36] LAI Y C,HUANG S W,YU H T.Gene structure and sequence polymorphism of the coat color gene,MC1R,in the Black-bellied vole (Eothenomys melanogaster)[J]. Zoological Studies,2016,55:e26.
[37] MORO O,IDETA R,IFUKU O.Characterization of the promoter region of the human melanocortin-1 receptor (MC1R) gene[J].Biochemical and Biophysical Research Communications,1999,262(2):452-460.
[38] 郭敏.狐狸毛色基因TYR、CBD103、MITF启动子活性探究[D].秦皇岛:河北科技师范学院,2018. GUO M.Study on promoter activity of fox fur gene TYR,CBD103,and MITF[D].Qinhuangdao:Hebei Normal University of Science and Technology,2018.(in Chinese)

Identification of the Core Promoter Region of MC1R Gene in Silver Fox

PDF (PC)

Knowledge

Abstract

Cite this article

share this article

References

Related Articles 15

Recommended Articles

Metrics

Comments

[1]	SONG Xingchao, MENG Jinzhu, ZHAO Yuanyuan, WU Zhenyang, AN Qingming. Cloning and Bioinformatics Analysis of MyoG Gene Promoter Region Sequence in Guizhou White Goat [J]. China Animal Husbandry and Veterinary Medicine, 2023, 50(3): 893-903.
[2]	GAO Xiaomin, ZHOU Shujian, CHEN Chen, JIN Jing, HU Cai, ZHANG Chen, ZUO Qisheng, ZHANG Yani, CHEN Guohong, LI Bichun. Regulation of STAT1 and Histone Acetylation Modification on lncRNA-BMP4 Transcription in Chickens [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(9): 3321-3332.
[3]	YANG Yuying, WANG Shaolin. Establishment of the Epidemiological Cut-off Values of Enterococcus Isolated from Animals for Antimicrobial Growth Promoters [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(7): 2788-2795.
[4]	ZHAI Zhe, CHEN Si, WU Yanru, WANG Xuemei, LI Chongrui, LIU Zhiyong, CHEN Qiaoling, WANG Fengyang, DU Li, LI Chang, JIN Ningyi. Analysis of Promoter Activity and Screening of Transcription Regulatory Elements of CCL19 Gene in Sheep [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(4): 1213-1222.
[5]	CHEN Zhe, YANG Pengxia, LEI Mingming, CHEN Jiabin, YAN Leyan. Cloning, Promoter Analysis and Transcriptional Regulation of p21 Gene in Goose [J]. China Animal Husbandry and Veterinary Medicine, 2022, 49(2): 405-415.
[6]	LU Xingrong, DUAN Anqin, MA Xiaoya, LIANG Shasha, DENG Tingxian. Construction and Analysis of the Luciferase Reporter Gene Vector of HSD17B1 Gene Promoter in Buffalo [J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(10): 3512-3521.
[7]	YANG Qin, DENG Xiaoyu, ZHAO Xiaoli, ZHANG Huan, XI Jing, YI Jihai, WANG Yong, ZHANG Qian, WANG Zhen, CHEN Chuangfu. Effects of Brucella Type Ⅳ Secretion System on Autophagy,Intracellular Survival and Inflammatory Response of Dendritic Cells [J]. China Animal Husbandry and Veterinary Medicine, 2021, 48(1): 257-264.
[8]	AI Jinxin, LONG Anju, LUO Weixing, CAI Huifen. Polymorphism and Bioinformatics Analysis of BMPR-ⅠB Gene Promoter Region in Guizhou Local Goats [J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(4): 1000-1008.
[9]	CHEN Wei, XU Houqiang, CHEN Xiang, QIN Hai, WU Yu, HUANG Mingjie, LU Xianjun. Promoters Activity Analysis of UCP3 and MYH1 Genes of Cattle in C2C12 and 3T3-L1 Cells [J]. China Animal Husbandry and Veterinary Medicine, 2020, 47(12): 3825-3832.
[10]	ZHANG Dongjie, WANG Liang, LIU Yang, LIU Di. Screening and Analysis of Transcription Factors Regulating Tβ4 Gene in Min Pig [J]. China Animal Husbandry and Veterinary Medicine, 2019, 46(9): 2535-2542.
[11]	ZHANG Feng, LI Xin, CHEN Mingxin. Transcriptional Regulation Analysis of Porcine SEPW1 Gene [J]. , 2019, 46(6): 1730-1738.
[12]	CHAO Zhe, WU Wangjun, XING Manping, HUANG Lili, SUN Ruiping, LIU Hailong, ZHENG Xinli. Detection of Single Nucleotide Polymorphism and Methylation of CpG Island in Promoter Region of Chicken HSP90AA1 Gene [J]. , 2019, 46(2): 504-511.
[13]	WANG Fei, ZONG Qiufang, MU Jingsheng, WU Shenglong, BAO Wenbin. Polymorphism and Bioinformatics Analysis of Promoter Regions of MYOD1 and AKT3 Genes in Large White Pigs [J]. China Animal Husbandry and Veterinary Medicine, 2019, 46(12): 3486-3494.
[14]	WANG Jingjing, YANG Hanyulu, DAI Rong, YANG Hua, SHI Guoqing, YANG Yonglin, WAN Pengcheng. Study on the Correlation Between HGTP Gene and Wool Fineness and the Activity of HGTP Gene Promoter [J]. China Animal Husbandry and Veterinary Medicine, 2019, 46(10): 2988-2997.
[15]	MAO Ning, WANG Jiafu, ZHANG Fuping, TANG Liangting, RUAN Yiqi, YI Fanli, RAN Xueqin. Study on Differential Expression of StAR and CYP11A1 Genes in Xiang Pig Ovary [J]. , 2018, 45(5): 1137-1144.