从江香猪基因组甲基化修饰及SNP与生长性状的关联研究

发布时间：2018-11-21 20:11

【摘要】：从江香猪是我国稀有的优良地方微型猪种,但香猪的生长慢的调控机制不清楚。本文利用Illumina Porcine SNP60(60K)猪全基因组SNP芯片对从江香猪生长性状关联SNP位点进行全基因组关联分析(GWAS),并结合MSAP检测从核基因整体水平分析从江香猪的遗传多态性,得到以下结论:1.应用甲基化敏感扩增多态性(MSAP)技术,通过筛选的10对引物对60头从江香猪个体血液基因组和肝脏DNA胞嘧啶甲基化模式和水平进行评估,克隆差异片段比对分析。总共得到5642条条带,从江香猪DNA的总甲基化水平为34.17%,全甲基化水平为14.84%,半甲基化水平为19.33%。分离克隆得到3条高体重群特有和3条低体重群特有的甲基化条带。其中A1-A5位于基因CLSTN3(Calsyntenin-3)、磷酸酰肌醇蛋白聚糖6(glypican-6-like)、泛素连接酶E3(HERC3)、蛋白酪氨酸磷酸酶受体G(Protein-tyrosine phosphatase gamma,PTPRG)内含子上,A6位于猪的UDP2C1基因第一个外显子前端(GeneID:LOC100515222,序列号NC_010450)。2.应用Illumina Porcine SNP 60芯片对120份香猪和可乐猪基因组的SNP进行检测,经过严格的质量控制,结合体长性状进行GWAS分析,得到15个与体长关联的SNP,挑选其中3个SNP位点INRA0011611、MARC0076145和H3GA0051838进行AS-PCR验证,结果显示芯片检测结果与AS-PCR检测结果相符。3.对3个生长相关SNP位点对12月龄香猪98份,18月龄香猪123份,24月龄香猪96份,大白猪48份,关岭猪18份,荣昌猪48份、糯谷猪30份、黔北黑猪23份、江口萝卜猪18份和宗地花猪28份样本进行AS-PCR检测。结果显示:位点1在各猪群体中基因型无偏好。大样本关联分析显示该点与24月龄香猪头长呈弱的正相关(0.2870.200)。位点2位于基因ALDH5A1第7内含子中,香猪群体中主要以G等位基因为优势等位基因,同时关联分析发现该点与18月龄香猪和24月龄香猪头长呈弱的负相关(-0.201-0.200,-0.310-0.200)。位点3位于基因TCEAL4上游,大样本验证结果显示在香猪群体中以GG基因型为主,G等位基因为优势等位基因。这些SNP位点功能的进一步研究和了解,对保护从江香猪种质资源,提高香猪养殖业发展速度具有一定借鉴作用。
[Abstract]:Congjiang Xiang pig is a rare and fine local miniature pig breed in China, but the regulation mechanism of its slow growth is not clear. In this paper, Illumina Porcine SNP60 (60K) porcine whole genome SNP chip was used to analyze the whole genome association SNP locus of Congjiang Xiang pig, and the genetic polymorphism of Congjiang Xiang pig was analyzed from the whole nuclear gene level with MSAP detection. The conclusions are as follows: 1. The methylation-sensitive amplified polymorphic (MSAP) (MSAP) technique was used to evaluate the methylation patterns and levels of DNA cytosine methylation in the blood and liver of 60 Congjiang pigs by screening 10 pairs of primers. A total of 5642 bands were obtained. The total methylation level of Congangxiang pig DNA was 34.17, the total methylation level was 14.84, and the semi-methylation level was 19.33. Three methylation bands specific to high body weight group and three low body weight groups were isolated and cloned. A1-A5 is located in intron of gene CLSTN3 (Calsyntenin-3), phosphoinositide proteoglycan 6 (glypican-6-like), ubiquitin ligase E3 (HERC3) and protein tyrosine phosphatase receptor G (Protein-tyrosine phosphatase gamma,PTPRG). A6 is located at the front end of the first exon of UDP2C1 gene (GeneID:LOC100515222, sequence number NC_010450). The genomic SNP of 120 fragrant pigs and cola pigs were detected by Illumina Porcine SNP 60 microarray. After strict quality control, the combined length traits were analyzed by GWAS, and 15 SNP, associated with body length were obtained, among which 3 SNP loci INRA0011611, were selected. MARC0076145 and H3GA0051838 are verified by AS-PCR. The results show that the results of chip detection are consistent with those of AS-PCR. 3. 3. Among them, 98 were 12 months old, 123 were 18 months old, 96 were 24 months old, 48 were big white pigs, 18 were Guan Ling pigs, 48 were Rongchang pigs, 30 were glutinous pigs, 23 were northern Guizhou black pigs. AS-PCR was performed on 18 radish pigs in Jiangkou and 28 in Zongduhua pigs. The results showed that locus 1 had no preference for genotype in all pig populations. Large sample correlation analysis showed that there was a weak positive correlation (0.2870.200) between this point and the head length of 24 months old fragrant pig. Locus 2 was located in intron 7 of gene ALDH5A1, and G allele was the dominant allele in Xiang pig population, and the correlation analysis showed that there was a weak negative correlation between G allele and head length (-0.201-0.200) in 18 month old Xiang pig and 24 month old Xiang pig. -0.310-0.200) Locus 3 was located upstream of gene TCEAL4. The results of large sample validation showed that GG genotype was dominant in Xiang pig population, and G allele was the dominant allele. The further study and understanding of the functions of these SNP loci can be used for reference in protecting the germplasm resources of Congjiang Xiang pigs and improving the development speed of Xiang pig breeding.
【学位授予单位】：贵州大学
【学位级别】：硕士
【学位授予年份】：2015
【分类号】：S828

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