摘要
【目的】克隆鉴定斑马鱼(Danio rerio)黑素皮质激素受体-1(Melanocortin-1 receptor,MC1R)基因编码区(Coding sequence,CDS),并进行生物信息学及序列变异分析,为开展MC1R基因生物学功能研究及揭示斑马鱼体色性状的调控机制提供基础资料。【方法】以红色、黄色和蓝色斑马鱼群体共计219个样本的肌肉组织DNA为模板,通过PCR扩增、克隆测序和拼接获得斑马鱼MC1R基因CDS序列,随后进行生物信息学分析和基因序列变异分析。【结果】获得MC1R基因全长1404 bp,CDS序列长972 bp,共编码323个氨基酸残基。MC1R蛋白含有7个跨膜结构域、4个胞外区和4个胞内区,呈典型的细胞表面受体样结构,主要定位于细胞质膜,存在4个潜在N-糖基化位点和26个磷酸化位点。斑马鱼MC1R基因与团头鲂和鱇浪白鱼的相似性较高,遗传距离最近,亲缘关系较密切。红色、黄色和蓝色斑马鱼群体共鉴定到2个变异位点,其中1个变异位点(g.334T>C)位于CDS,存在TT和TC基因型,该变异未引起氨基酸的改变,另1个变异位点位于3'-非编码区(3'-UTR)(g.276A>T),存在AA和AT基因型,且不同体色间2个变异位点g.334T>C和g.276A>T的基因型频率均一致。【结论】MC1R基因序列在不同鱼类中既具保守性,又存在一定差异,蛋白结构在不同鱼类中可能相对保守且功能类似。MC1R基因可作为开展不同鱼类系统进化关系研究的分子标记,其在红色、黄色和蓝色斑马鱼群体中较保守,存在g.276A>T和g.334T>C变异位点,且2个变异位点可能存在连锁关系。
【Objective】This study aimed to clone and identify the coding sequence(CDS)of the melanocortin-1 re-ceptor(MC1R)gene in zebrafish(Danio rerio),and to conduct bioinformatics and sequence variation analysis,so as to provide fundamental materials for elucidating the biological function of the MC1R gene and the regulation mechanisms of zebrafish body color traits.【Method】Using muscle tissue DNA from 219 samples of red,yellow,and blue zebrafish populations as templates,CDS sequences of the MC1R gene of zebrafish were obtained through PCR amplification,clo-ning,sequencing,and assembly to perform bioinformatics analysis and gene sequence variation analysis.【Result】The full-length of MC1R gene was 1404 bp,with a CDS of 972 bp,encoding 323 amino acid residues.The MC1R protein con-tained seven transmembrane domains,four extracellular regions,and four intracellular regions,exhibiting a typical cell surface receptor-like structure.The protein was mainly localized to the plasma membrane and possessed four potential N-glycosylation sites and 26 phosphorylation sites.The zebrafish MC1R gene showed high similarity to that of Megalobrama amblycephala and Anabarilius grahami,demonstrating the nearest genetic distance and close genetic relationships.Two variant sites were identified in red,yellow,and blue zebrafish populations:one variant site(g.334T>C)was located in the CDS and exhibited both TT and TC genotypes,without causing any change in amino acid;the other variant site was located in the 3'-untranslated region(3'-UTR)(g.276A>T)and exhibited both AA and AT genotypes.The frequencies of the two variants,g.334T>C and g.276A>T,were consistent across color groups.【Conclusion】The MC1R gene sequence is conserved and somewhat diverse across fish species,suggesting that the protein structure is relatively conserved and functionally similar across species.The MC1R gene can be used as an effective molecular marker for studying the evolu-tionary relationships of different fish systems.It is relatively conserved in red,yellow,and blue zebrafish populations,and contains two variant sites(g.276A>T and g.334T>C)which may be in linkage.
作者
宋兴超
王莎莎
王云艳
梅杰
梁正其
巴家文
SONG Xing-chao;WANG Sha-sha;WANG Yun-yan;MEI Jie;LIANG Zheng-qi;BA Jia-wen(College of Agroforesry Engineering and Planning,Tongren University/Guizhou Provincial Key Laboratory for Biodiver‐sity Conservation and Utilization in the Fanjing Mountain Region,Tongren,Guizhou 554300,China)
出处
《南方农业学报》
北大核心
2025年第7期2317-2327,共11页
Journal of Southern Agriculture
基金
国家自然科学基金项目(32060274)
贵州省科技计划项目(黔科合基础-ZK〔2022〕一般559)
贵州省高层次创新型(千层次)人才项目(2024-〔2022〕-048)
贵州省大学生创新创业训练计划项目(S2024106651872)。
作者简介
通信作者:巴家文(1984-),https://orcid.org/0009-0002-3468-065X,博士,教授,主要从事鱼类生态学研究工作,E-mail:bajiawen@163.com;第一作者:宋兴超(1982-),https://orcid.org/0000-0001-5484-9526,博士,副教授,主要从事水产动物遗传育种研究工作,E-mail:songxingchao_888@126.com。
