赤水乌骨鸡EDNRB2基因多态性及其与肤色性状的关联分析

doi:10.16431/j.cnki.1671-7236.2025.02.026

摘要/Abstract

摘要： 【目的】分析赤水乌骨鸡内皮素受体B亚型2(endothelin receptor B subtype 2，EDNRB2)基因多态性对乌骨鸡肤色性状的影响，并筛选EDNRB2基因调控赤水乌骨鸡肤色的关键多态位点，为深入探究乌骨鸡黑色素沉积调控的内在机制与赤水乌骨鸡肤色性状的优化改良提供新的遗传标记。【方法】以赤水乌骨鸡作为研究对象，对EDNRB2基因第1～9外显子进行PCR扩增后测序，对所获序列进行比对鉴定其单核苷酸多态性(SNP)位点，并分析EDNRB2基因的遗传多样性。利用RNAfold web server分析EDNRB2基因SNP位点突变前后的mRNA二级结构；利用SHEsis在线网站进行EDNRB2基因SNP位点的连锁不平衡及单倍型、双倍型分析；利用SPSS 27.0软件分析EDNRB2基因多态性与赤水乌骨鸡肤色性状的关联性。【结果】在赤水乌骨鸡EDNRB2基因第3与9外显子上共发现5个SNPs位点，其中第3外显子中存在2个SNPs位点(g.11120193 T＞C和g.11120382 C＞T)，第9外显子中在3个SNPs位点(g.11125882 G＞A、g.11125917 T＞A、g.11125931 C＞A)。g.11120193 T＞C位点存在TT、TC、CC 3种基因型，g.11120382 C＞T位点存在CC、CT 2种基因型，突变类型均属于同义突变；g.11125882 G＞A位点存在GG、AG、AA 3种基因型，属于错义突变(Gly→Glu)；g.11125917 T＞A位点存在TT、AT、AA 3种基因型，属于错义突变(Phe→Ile)；g.11125931 C＞A位点存在CC、AC、AA 3种基因型，属于无义突变。EDNRB2基因5个SNPs位点共发现10种单倍型与24种双倍型，其中优势单倍型与双倍型分别为H4与H4H4。关联分析结果显示，EDNRB2基因g.11120193 T＞C位点CC基因型个体的鸡冠△a值显著高于TC基因型(P＜0.05)，背部△a值显著低于TT基因型(P＜0.05)；g.11125931 C＞A位点CC基因型个体鸡冠△L值显著高于AC基因型(P＜0.05)；其余SNP位点不同基因型个体间的肤色参数差异均不显著(P＞0.05)。双倍型H5H5与H7H7对赤水乌骨鸡的肤色性状影响最为明显。【结论】在EDNRB2基因第3、9外显子上共发现5个SNPs位点，其中g.11120193 T＞C和g.11125931 C＞A位点与赤水乌骨鸡的鸡冠、背部的肤色有显著关联性。双倍型H5H5对鸡冠△L值与△a值、背部△b值、翅下△b值有明显影响。EDNRB2基因能够作为赤水乌骨鸡的肤色性状育种选择的候选基因。

关键词: 赤水乌骨鸡; EDNRB2基因; 单核苷酸多态性; 肤色; 关联分析

Abstract: 【Objective】 This study was aimed to analyze the effects of polymorphisms in endothelin receptor B subtype 2 (EDNRB2) gene on skin color traits in Chishui Black-bone chickens,and screen the key polymorphism loci of EDNRB2 gene in regulating skin color of Chishui Black-bone chickens,in order to provide new genetic markers for a thorough examination of the intrinsic mechanism regulating melanin deposition in silky fowls and the optimization and improvement of skin color traits in Chishui Black-bone chickens. 【Method】 Chishui Black-bone chickens were used as the research subject in this study.The nine exons of EDNRB2 gene were sequenced after PCR amplification,the single nucleotide polymorphism (SNP) of EDNRB2 gene were identified by comparison,and the genetic diversity of EDNRB2 gene were analyzed.The mRNA secondary structure before and after the mutation of EDNRB2 gene SNPs was analyzed by RNAfold web server.The chain disequilibrium,haplotype and diplotype of EDNRB2 gene SNP were analyzed using SHEsis online website.The correlation between EDNRB2 gene SNP and skin color traits in Chishui Black-bone chickens was analyzed by SPSS 27.0 software. 【Result】 Five SNPs were identified in exons 3 and 9 of EDNRB2 gene in Chishui Black-bone chickens,of which two SNPs of g.11120193 T＞C and g.11120382 C＞T were found to exist in exon 3,and of which three SNPs of g.11125882 G＞A,g.11125917 T＞A and g.11125931 C＞A were found to exist in exon 9.There were three genotypes of TT,TC and CC in g.11120193 T＞C,and there were two genotypes of CC and CT in g.11120382 C＞T,which belonged to samesense mutation.There were three genotypes of GG,AG and AA in g.11125882 G＞A,which belonged to missense mutation (Gly→Glu).There were three genotypes of TT,AT and AA in g.11125917 T＞A,which belonged to missense mutation (Phe→Ile).There were three genotypes of CC,AC and AA in g.11125931 C＞A,which belonged to nonsense mutation.Five SNPs of EDNRB2 gene had a total of 10 haplotypes and 24 diplotypes,with H4 and H4H4 being the dominant haplotype and diplotype,respectively.The results of association analysis showed that the cockscomb △a value of CC genotype of EDNRB2 gene in g.11120193 T＞C was significantly higher than that of TC genotype (P＜0.05),the back △a value was significantly lower than that of TT genotype (P＜0.05).The cockscomb △L value of CC genotype of EDNRB2 gene in g.11125931 C＞A was significantly higher than that of AC genotype (P＜0.05).At the remaining SNPs,there were no significant variations in skin color parameters across individuals with different genotypes (P＞0.05).The most noticeable effects on the skin color traits in Chishui Black-bone chickens were caused by haplotypes H5H5 and H7H7. 【Conclusion】 EDNRB2 gene had five SNPs in exons 3 and 9,with g.11120193 T＞C and g.11125931 C＞A showing a substantial correlation with the skin color of cockscomb and back in Chishui Black-bone chickens.The haplotype H5H5 significantly impacted the cockscomb △L and △a values,back △b value,and underwing △b value.EDNRB2 gene could be utilized as a potential gene for breeding selection of skin color traits in Chishui Black-bone chickens.

Key words: Chishui Black-bone chickens; EDNRB2 gene; single nucleotide polymorphism; skin color; association analysis

中图分类号:

S813.3

杨金林, 李辉, 赵德鹏, 石钰仕, 龙霞, 谭启松. 赤水乌骨鸡EDNRB2基因多态性及其与肤色性状的关联分析[J]. 中国畜牧兽医, 2025, 52(2): 759-770.

YANG Jinlin, LI Hui, ZHAO Depeng, SHI Yushi, LONG Xia, TAN Qisong. Polymorphisms of EDNRB2 Gene and Its Association with Skin Color Traits in Chishui Black-bone Chickens[J]. China Animal Husbandry and Veterinary Medicine, 2025, 52(2): 759-770.

参考文献

[1] 叶涛.鸡SLCO1B3基因多态性研究及其互作蛋白筛选分析[D].贵阳:贵州大学,2022.YE T.Polymorphism of chicken SLCO1B3 gene and its interaction protein screening analysis[D].Guiyang:Guizhou University,2022.(in Chinese)
[2] 李思.赤水乌骨鸡种质特性及其黑色素保健价值的研究[D].贵阳:贵州大学,2015.LI S.Study on germplasm characteristics of Chishui Black-bone chickens and the value of health care of its melanin[D].Guiyang:Guizhou University,2015.(in Chinese)
[3] KAWASAKI-NISHIHARA A,NISHIHARA D,NAKAMURA H,et al.ET3/EDNRB2 signaling is critically involved in regulating melanophore migration in Xenopus[J]. Developmental Dynamics,2011,240(6):1454-1466.
[4] LIU H,LUO Q,ZHANG J,et al.Endothelins (EDN1,EDN2,EDN3) and their receptors (EDNRA,EDNRB,EDNRB2) in chickens:Functional analysis and tissue distribution[J]. General and Comparative Endocrinology,2019,283:113231.
[5] LI D,WANG X,FU Y,et al.Transcriptome analysis of the breast muscle of Xichuan Black-bone chickens under tyrosine supplementation revealed the mechanism of tyrosine-induced melanin deposition[J].Frontiers in Genetics,2019,10:457.
[6] ZI X,GE X,ZHU Y,et al.Transcriptome profile analysis identifies candidate genes for the melanin pigmentation of skin in Tengchong Snow chickens[J].Veterinary Sciences,2023,10(5)：341.
[7] YANG Y,WANG C,LIU Y,et al.Comparative transcriptomic analyses reveal key genes underlying melanin distribution during embryonic development in geese (Anser anser)[J]. British Poultry Science,2024,65(4):387-393.
[8] MAO N N,WANG W J,ZHANG R,et al.Population genomics reveals that a missense mutation in EDNRB2 contributes to white plumage color in pigeons[J].Poultry Science,2024,103(1):103225.
[9] MACLARY E T,WAUER R,PHILLIPS B,et al.An allelic series at the EDNRB2 locus controls diverse piebalding patterns in the domestic pigeon[J].PLoS Genetics,2023,19(10):e1010880.
[10] YANG Y,WANG H,LI G,et al.Using comparative genomics to detect mutations regulating plumage variations in Graylag (A.anser) and Swan geese (A.cygnoides)[J].Gene,2022,834:146612.
[11] GUO Q,JIANG Y,WANG Z,et al.Genome-wide association study for screening and identifying potential shin color loci in ducks[J].Genes,2022,13(8)：1391.
[12] GUO Q,JIANG Y,WANG Z,et al.Genome-wide analysis identifies candidate genes encoding beak color of duck[J].Genes,2022,13(7)：1271.
[13] MACLARY E T,PHILLIPS B,WAUER R,et al.Two genomic loci control three eye colors in the domestic pigeon (Columba livia)[J].Molecular Biology and Evolution,2021,38(12):5376-5390.
[14] KINOSHITA K,AKIYAMA T,MIZUTANI M,et al.Endothelin receptor B2(EDNRB2) is responsible for the tyrosinase-independent recessive white (mo(w)) and mottled (mo) plumage phenotypes in the chicken[J].PLoS One,2014,9(1):e86361.
[15] 刘航,王欢欢,葛莹,等.基于转录组和蛋白组筛选乌骨鸡肤色性状候选基因[J].畜牧兽医学报,2023,54(6):2320-2329.LIU H,WANG H H,GE Y,et al.Screening of candidate genes of skin color of Black-bone chicken based on transcriptome and proteome[J].Acta Veterinaria et Zootechnica Sinica,2023,54(6):2320-2329.(in Chinese)
[16] XU M,TANG S,LIU X,et al.Genes influencing deposition of melanin in breast muscle of the Xuefeng Black bone chicken based on bioinformatic analysis[J].Genome,2023,66(8):212-223.
[17] WU N,QIN H,WANG M,et al.Variations in endothelin receptor B subtype 2(EDNRB2) coding sequences and mRNA expression levels in 4 Muscovy duck plumage colour phenotypes[J].British Poultry Science,2017,58(2):116-121.
[18] MIWA M,INOUE-MURAYAMA M,AOKI H,et al.Endothelin receptor B2(EDNRB2) is associated with the panda plumage colour mutation in Japanese quail[J].Animal Genetics,2007,38(2):103-108.
[19] 屠云洁,栾德琴,章明,等.ALDH7A1和EDNRB2基因分型及其与乌骨鸡皮肤乌色度关联分析[J].中国畜牧兽医,2024,51(7):2923-2932.TU Y J,LUAN D Q,ZHANG M,et al.Genotyping of ALDH7A1 and EDNRB2 genes and their association with skin blackness in Black-bone chickens[J].China Animal Husbandry & Veterinary Medicine,2024，51(7):2923-2932.(in Chinese)
[20] 叶涛,李辉,罗韦,等.赤水乌骨鸡SLCO1B3基因SNPs鉴定及与蛋品质的关联性分析[J].畜牧与兽医,2023,55(2):25-31.YE T,LI H,LUO W,et al.SNPs identification of the SLCO1B3 gene and its correlation with the egg quality of Chishui Black-bone chicken[J].Animal Husbandry & Veterinary Medicine,2023,55(2):25-31.(in Chinese)
[21] HARRIS M L,HALL R,ERICKSON C A.Directing pathfinding along the dorsolateral path—The role of EDNRB2 and EPHB2 in overcoming inhibition[J].Development,2008,135(24):4113-4122.
[22] PLA P,ALBERTI C,SOLOV’EVA O,et al.EDNRB2 orients cell migration towards the dorsolateral neural crest pathway and promotes melanocyte differentiation[J].Pigment Cell Research,2005,18(3):181-187.
[23] 王琛.mRNA二级结构自由能在不同物种中的分布差异研究[D].杨凌:西北农林科技大学,2015.WANG C.Studies on the difference of mRNA secondary structure and MFE among different species[D].Yangling:Northwest A&F University,2015.(in Chinese)
[24] 毛圆辉.翻译过程中mRNA二级结构的功能研究[D].杨凌:西北农林科技大学,2015.MAO Y H.A systematic analysis of roles of mRNA secondary structures during translation[D].Yangling:Northwest A&F University,2015.(in Chinese)
[25] LIU C,ZOU Q,TANG H,et al.Melanin nanoparticles alleviate sepsis-induced myocardial injury by suppressing ferroptosis and inflammation[J].Bioactive Materials,2023,24:313-321.
[26] GUO K,JIAO Z,ZHAO X,et al.Melanin-based immunoregulatory nanohybrids enhance antitumor immune responses in breast cancer mouse model[J].ACS Nano,2023,17(11):10792-10805.
[27] 杨亚杰,李慧,胡清清,等.乌骨鸡黑色素的体外抗氧化活性及其对人脐静脉内皮细胞氧化损伤的保护作用[J].南昌大学学报(理科版),2019,43(1):59-64.YANG Y J,LI H,HU Q Q,et al.Protective effect of melanin from Black-bone Sliky fowl on human umbilical vein endothelial cell injury and antioxidant activity in vitro[J].Journal of Nanchang University(Natural Science), 2019,43(1):59-64.(in Chinese)