牦牛TMEM-18基因多态性及其与生产性能关联分析

doi:10.13560/j.cnki.biotech.bull.1985.2017.02.013

摘要/Abstract

摘要： 跨膜蛋白18基因（TMEM-18）是一个末端为低聚嘧啶的基因,与动物生长发育密切相关。为了研究牦牛TMEM-18 基因多态性与生产性能的关系。以192头天祝雌性牦牛血样NDA构建混合池,扩增TMEM-18基因内含子1和外显子5的序列。运用BLAST和Chromas软件分析突变位点,运用高分辨率熔解曲线分析技术（HRM）统计基因型分型,采用SHEsis 软件进行基因型频率和等位基因频率计算,同时对多态位点进行配对连锁不平衡和单倍型分析,运用SPSS21.0分析基因多态位点与生产性能关联。结果表明,牦牛TMEM-18基因内含子1处存在2个多态位点,分别是861（A/G）和1267（C/T）,外显子5处存在1个多态位点为4 447（C/T）。关联分析表明,牦牛TMEM-18基因861（A/G）位点的不同基因型与牦牛的体高、体重、胴体重和屠宰率差异性显著（P<0.05）;1267（C/T）位点的不同基因型与牦牛的体高、体重和屠宰率差异性显著（P<0.05）,而牦牛TMEM-18基因4447（C/T）位点的不同基因型与牦牛的体斜长、管围、胸围、体重、胴体重、净肉重、净肉率和屠宰率均存在显著性差异（P<0.05）。通过单倍型分析发现群体中存在3种单倍型组合,即ACC单倍型、GTC单倍型和GTT单倍型,其中ACC单倍型组合个体数目明显多于其他单倍型组合,为优势单倍型。本实验揭示TMEM-18基因可作为牦牛生产性能开发的候选分子标记,为牦牛遗传资源的利用、开发与新品种的选育提供依据。

关键词: 牦牛, TMEM-18基因, SNP, HRM, 生产性状

Abstract: The TMEM-18（transmembrane protein 18）gene is the one with oligomer in its terminal,and closely related to the growth and development of animal. This study aims to study the correlation between polymorphism of TMEM-18gene and production traits in yak. Firstly,constructing DNA mixing pool with blood samples from 192 female Tianzhu yaks,and the intron1 and exon5 of TMEM-18 gene were amplified. Then,BLAST and Chromas were used to analyze the mutation sites in TMEM-18 gene,high-resolution melting curve（HRM）to determine the genotype,software SHEsis to calculate and the frequencies of genotypes and alleles and to analyze the linkage disequilibrium and haplotypes in the SNPs of TMEM-18 gene,and software SPSS21.0 to understand the correlation between gene polymorphic loci and product traits. The experimental results showed that the intron1 of yak TMEM-18 gene contained 2 polymorphic loci of 861（A/G）and 1 267（C/T）,and the exon5 contained 1 polymorphic locus of 4 447（C/T）. Correlation analysis indicated that the different genotypes at locus 861（A/G）resulted in the significant differences on body height,body weight,carcass weight,and slaughter rate（P<0.05）;while the varied genotypes at locus 1 267（C/T）caused significant differences on body height,body weight and slaughter rate（P<0.05）;and the various genotypes at locus 4 447（C/T）let to significant differences on body length,tube girth,chest girth,body weight,carcass weight,meat weight,net meat rate,and slaughter rate（P<0.05）. There were 3 haplotype combinations in the yak populations,namely ACC,GTC and GTT. Among them,the number of ACC was significantly more than that of others,i.e.,it was dominant haplotype. The results suggested that the TMEM-18 gene could be used as molecular-marker candidate for developing the production traits of yak,providing a scientific basis for the utilization and development of yak genetic resources and breeding of new varieties.

Key words: yak, TMEM-18 gene, SNP, HRM, production traits

张欢, 张全伟, 王琪, 马友记, 张勇, 赵兴绪. 牦牛TMEM-18基因多态性及其与生产性能关联分析[J]. 生物技术通报, 2017, 33(2): 89-96.

ZHANG Huan, ZHANG Quan-wei, WANG Qi, MA You-ji, ZHANG Yong, ZHAO Xing-xu. Polymorphism of TMEM-18 Gene and Its Correlation with Production Traits in Yak[J]. Biotechnology Bulletin, 2017, 33(2): 89-96.

参考文献

[1] 何梅兰, 马秀山. 天祝白牦牛产业发展现状及发展战略[J]. 中国草食动物科学, 2015, 35（5）：71-74.
[2] 马苍. 天祝白牦牛产业可持续发展思考[J]. 家畜生态学报, 2008, 29（6）：133-135.
[3] 梁春年. 牦牛MSTN和IGF-IR基因的克隆及SNPs与生长性状相关性[D]. 兰州：甘肃农业大学, 2011.
[4] 李亚兰. 天祝白牦牛IGF-2基因的全长cDNA克隆, 组织表达及生产性状相关性分析[D]. 兰州：甘肃农业大学, 2015.
[5] 李天科. 牦牛GDF-10, IHH, MC4R基因的多态与生长性状的相关性研究[D]. 兰州：甘肃农业大学, 2014.
[6] Jurvansuu J, Zhao Y, Leung DS, et al. Transmembrane protein 18 enhances the tropism of neural stem cells for glioma cells[J]. Cancer Research, 2008, 68（12）：4614-4622.
[7] Hindorff LA, Sethupathy P, Junkins HA, et al. Potential etiologic and functional implications of genome-wide association loci for human diseases and traits[J]. Proc Natl Acad Sci USA, 2009, 106（23）：9362-9367.
[8] Gutierrez-Aguilar R, Kim DH, Woods SC, et al. Expression of new loci associated with obesity in diet-induced obese rats：from genetics to physiology[J]. Obesity（Silver Spring）, 2011, 20, 306-312.
[9] Bernhard F, Landgraf K, Klöting N, et al. Functional relevance of genes implicated by obesity genome wide association study signals for human adipocyte biology[J]. Diabetologia, 2013, 56（2）：311-322.
[10] Rask-Andersen M, Jacobsson JA, Moschonis G, et al. Association of TMEM18 variants with BMI and waist circumference in children and correlation of mRNA expression in the PFC with body weight in rats[J]. European Journal of Human Genetics, 2012, 20（2）：192-197.
[11] Hotta K, Nakamura M, Nakamura T, et al. Association between obesity and polymorphisms in SEC16B, TMEM18, GNPDA2, BDNF, FAIM2 and MC4R in a Japanese population[J]. Journal of Human Genetics, 2009, 54（12）：727-731.
[12] Holzapfel C, Grallert H, Huth C, et al. Genes and lifestyle factors in obesity：Results from 12462 subjects from MONICA/KORA[J]. International Journal of Obesity, 2010, 34（10）：1538-1545.
[13] Hong KW, Oh B. Recapitulation of genome-wide association studies on body mass index in the Korean population[J]. International Journal of Obesity, 2011, 36（8）：1127-1130.
[14] Zhao J, Bradfield JP, Li M, et al. The role of obesity-associated loci identified in genome-wide association studies in the determination of pediatric BMI[J]. Obesity（Silver Spring）, 2009, 17（12）：2254-2257.
[15] Almén MS, Jacobsson JA, Shaik JH, et al. The obesity gene, TMEM18, is of ancient origin, found in majority of neuronal cells in all major brain regions and associated with obesity in severely obese children[J]. BMC Medical Genetics, 2010, 11（1）：58.
[16] 林平. 大白猪和通城猪肌纤维组织学特性差异和SH2B1, TEM18, IDH3B基因遗传效应分析[D]. 武汉：华中农业大学, 2013.
[17] 马伟. 牛TMEM18基因克隆, SNP检测及其与部分经济性状的关联分析[D]. 西安：西北农林科技大学, 2012.
[18] 郭宪, 李天科, 裴杰, 阎萍等. 南德温杂交肉牛生长分化因子10基因多态性与体尺性状的关联分析[J]. 中国畜牧兽医, 2014, 41（12）：197-203.
[19] Michael L, Robert P, Robert P, et al. Genotyping of single-nucleotide polymorphisms by high- resolution melting of small amplicons[J]. Molecular Diagnostics and Genetics, 2004, 50（7）：1156-1164.
[20] Ye MH, Chen JL, Zhao GP, et al. Associations of A-FABP and H-FABP markers with the content of intramuscular-lar fat in Beijing-you chicken[J]. Animal Biotechnology, 2010, 21：14-24.
[21] 曾凯. 群体遗传学理论在检测正向选择与研究物种形成机制中的应用[D]. 广州：中山大学, 2007.
[22] Cameron N, Steven F. Base-pair neutral homozygotes can be discriminated by calibrated high-resolution melting of small amplicons[J]. Nucleic Acids Research, 2008, 1093（10）：1-8.
[23] Nei M, Roychoudhury AK. Sampling variances of heterqozygosity and genetic distance[J]. Genetics, 1974, 76（2）：379-390.
[24] Nei M, Li WH. Mathematical model for studying genetic variation in terms of restriction endonucleases[J]. Proc Natl Acad Sci USA, 1979, 76（10）：5269-5273.
[25] Yamashita R, Suzuki Y, Takeuchi N, et al. Comprehensive detection of human terminal oligo-pyrimidine（TOP）genes and analysis of their characteristics[J]. Nucleic Acids Research, 2008, 36（11）：3707-3715.
[26] 牛晓亮. 牦牛CAPN4和CAST基因多态性及与胴体和肉质性状关联研究[D]. 兰州：甘肃农业大学, 2015.
[27] Caws E, White SN, Riley DG, et al. Assessment of single nucleotide polymorphisms in genes residing on chromosomes 14 and 29 for association with carcass composition traits in Bos indicus cattle[J]. Journal of Animal Science, 2005, 83（1）：13-19.
[28] Chang KC. Critical regulatory domains in intron 2 of porcine sarcomeric myosin heavy chain gene[J]. Journal of Muscle Research and Cell Motility, 2000, 21：451-461.
[29] 张冉. 肉质性状相关基因的表达及消化道菌群多样性与动物生长发育的关系[D]. 济南：山东大学, 2014.