摘要
为筛选与中国荷斯坦牛繁殖和泌乳性状相关的遗传标记,本研究从分子水平探究了MET基因单核苷酸多态(single nucleotide polymorphisms,SNPs)与繁殖和泌乳性状的相关性。通过混池测序法在70头无血缘关系的健康中国荷斯坦公牛中扫描目标基因的SNPs,采用竞争性等位基因特异性PCR(kompetitive allele-specific PCR,KASP)对其后代1160头中国荷斯坦泌乳牛进行部分SNP的基因分型,基于连锁不平衡分析获得双倍型信息,并采用线性模型对SNP及其双倍型与5个繁殖和6个泌乳性状的估计育种值(estimated breeding values,EBV)进行关联分析。随后,针对位于功能区域的SNPs进行生物信息学分析和基因表达量相关性分析,初步探究其潜在的调控机制。本研究在MET基因中共检测到19个SNPs,说明该基因存在丰富的遗传变异;就其中7个SNPs进行了基因分型,分型结果与目标性状的关联分析结果显示,7个SNPs与多个性状存在显著(P<0.05)或极显著(P<0.01)的关联;位于上游调控区的g.51737889T>C位点GG基因型个体的初产日龄、青年牛首末配种间隔、经产牛首末配种间隔、体细胞评分的EBV最低,说明该基因型个体同时具有较好的繁殖性能和泌乳性能;另一个位于上游调控区的g.51736640A>C位点GG基因型个体的初产日龄、青年牛首末配种间隔、经产牛首末配种间隔、体细胞评分的EBV最低,但乳蛋白率、乳脂率的EBV最低,说明该基因型个体繁殖性能较好,但泌乳性能相对较差;此外,位于第6外显子的g.51660569G>A位点TC基因型个体初产日龄的EBV较低,青年牛首末配种间隔、经产牛首末配种间隔、体细胞评分的EBV最低,说明该基因型个体具有较好的繁殖性能。连锁不平衡分析发现,该基因的7个SNPs形成2个单倍型块,其双倍型与目标性状的关联分析进一步表明,单倍型块1中H1H3个体的繁殖性能较好且泌乳性能中等,单倍型块2中H4H4个体的繁殖性能和泌乳性能均较优秀,为两个优势双倍型;此外,H1H3包含g.51660569G>A位点的TC基因型,H4H4为g.51737889T>C、g.51736640A>C两位点GG基因型的组合,其关联分析结果与单个位点的结果一致。基序分析结果表明,g.51737889T>C与g.51736640A>C位点的优势等位基因G可富集到与基因激活、细胞增殖、分化相关的转录因子,且两位点GG基因型的MET基因表达量均最高,其双倍型H4H4表达量也较高,进一步验证了关联分析结果。综上,本研究获得了中国荷斯坦牛MET基因的多态性图谱,并通过关联分析、生物信息学预测及基因表达量分析发现并验证了MET基因与繁殖、泌乳性状的遗传关联,为中国荷斯坦牛高产高效选育提供了可用的遗传标记。
In order to identify genetic markers related to reproduction and milk production traits of Chinese Holstein cattle, the relationships between single nucleotide polymorphisms(SNPs) of MET gene and reproduction and milk production traits was explored. Candidate SNPs of MET were screened through direct sequencing of pooled DNA samples in 70 healthy unrelated Chinese Holstein bulls, and the genotyping of partial SNPs in 1 160 healthy Holstein lactating cattle was performed using kompetitive allele-specific PCR(KASP) method, diplotype information based on linkage disequilibrium analysis was obtained. The linear models were employed to conduct the association analysis between SNPs and their diplotypes and EBVs of 5 reproductive traits and 6 milk production traits. The bioinformatics analysis and the association analysis were conducted on SNPs located in functional areas. In this study, a total of 19 SNPs were detected in MET, indicating rich genetic variations existing within this gene in the population. Among them, 7 SNPs was genotyped, and the association analysis revealed that these SNPs were significantly(P<0.05) or highly significantly(P<0.01) associated with multiple traits. The individuals with GG genotype of g.51737889 T>C located in upstream had the lowest EBVs of the age at the first calving(AFC), the interval from the first to last insemination in heifer(IFL_H), the interval from the first to last insemination in cows(IFL_C), and somatic cell score(SCS), which suggested the good reproduction and milk production performance of animals carrying this genotype. The individuals with GG genotype of g.51736640 A>C located in upstream had the lowest EBVs of the age at the first calving, the interval from the first to last insemination in heifers, the interval from the first to last insemination in cows, and somatic cell score;however, they had the lowest EBVs of protein percentage(PP), and fat percentage(FP), which suggested animals carrying this genotype had the good performance in reproductive, but with relatively poor performance in milk production. The individuals with the TC genotype at g.51660569 G>A in 6exon had the lowest EBVs of the interval from the first to last insemination in heifers and the interval from the first to last insemination in cows, which suggested the good reproduction performance of animals carrying this genotype. Linkage disequilibrium analysis revealed that 7 SNPs linked to form 2 haplotype blocks. The association analysis between diplotypes and target traits showed that the individuals with H1 H3 diplotype at BLOCK1 had good reproduction performance but medium milk production performance, the individuals with H4 H4 diplotype at BLOCK2 had good reproduction and milk production performance, and these two were dominant hiplotypes. In addition, H1 H3 diplotype included the TC genotype at g.51660569 G>A, H4 H4 diplotype was combination of the GG genotype at g.51737889 T>C and g.51736640 A>C, the results of diplotype association analysis were consistent with that of SNP. The results of motif analysis showed that G allele at g.51737889 T>C and g.51736640 A>C enriched in transcription factors related to gene activation, cell proliferation and differentiation. The individuals with GG genotype at both SNPs had the highest gene expression, and the individuals with corresponding H4 H4 diplotype had higher gene expression, which further verified the results of association analysis. In conclusion, the polymorphism map of MET gene was derived in Chinese Holstein cattle, and the genetic association between MET gene and reproductive and productive traits was identified through association analysis, and verified by bioinformatics prediction and gene expression analysis. The results provided useful genetic markers for genetic selection of Chinese Holstein cattle for higher production as well as higher efficiency.
作者
许静漪
徐昊祺
胡丽蓉
张帆
高清
罗汉鹏
张海亮
师睿
李想
刘林
郭刚
王雅春
XU Jingyi;XU Haoqi;HU Lirong;ZHANG Fan;GAO Qing;LUO Hanpeng;ZHANG Hailiang;SHI Rui;LI Xiang;LIU Lin;GUO Gang;WANG Yachun(National Engineering Laboratory for Animal Breeding,Key Laboratory of Animal Genetics,Breeding and Reproductive of Ministry of Agriculture and Rural Affairs,College of Animal Science and Technology,China Agricultural University,Beijing 100193,China;College of Animal Science,Xinjiang Agricultural University,Urumqi 830052,China;Beijing Dairy Cattle Center,Beijing 100194,China;Beijing Sunlon Livestock Development Company Limited,Beijing 100029,China)
出处
《畜牧兽医学报》
CAS
CSCD
北大核心
2022年第11期3769-3785,共17页
ACTA VETERINARIA ET ZOOTECHNICA SINICA
基金
现代农业产业技术体系(CARS-36)
长江学者和创新团队发展计划(IRT15R62)
北京三元种业科技股份有限公司自立科研课题(SYZYZ20190005)。
关键词
MET
单核苷酸多态性
单倍型
基因表达
奶牛
关联分析
MET
single nucleotide polymorphism
haplotype
gene expression
cow
association analysis