中国农业科学 ›› 2016, Vol. 49 ›› Issue (4): 739-753.doi: 10.3864/j.issn.0578-1752.2016.04.013

• 畜牧·兽医·资源昆虫 • 上一篇    下一篇

杜长大猪、二花脸猪和莱芜猪3个群体25种血液性状的全基因组关联分析

刘晨龙,杨慧,张慧,张志燕,段艳宇   

  1. 江西农业大学省部共建猪遗传改良与养殖技术国家重点实验室,南昌 330045
  • 收稿日期:2014-10-02 出版日期:2016-02-16 发布日期:2016-02-16
  • 通讯作者: 段艳宇,E-mail:yanyuduan@gmail.com
  • 作者简介:刘晨龙,E-mail:liuchenlong0102@hotmail.com
  • 基金资助:
    国家“973”项目(2012CB722502)、甘肃省生物技术专项(31200926)、江西省科技支撑计划(2009BNA06900)

Genome-Wide Association Study of 25 Hematological Parameters in Western DLY Pigs and Two Chinese Erhualian and Laiwu Populations

LIU Chen-long, YANG Hui, ZHANG Hui, ZHANG Zhi-yan, DUAN Yan-yu   

  1. National Key Laboratory for Swine Genetics, Breeding and Production Technology, Jiangxi Agricultural University, Nanchang 330045
  • Received:2014-10-02 Online:2016-02-16 Published:2016-02-16

摘要: 【目的】 利用全基因组关联分析定位影响杜长大猪(DLY)、二花脸猪(EHL)和莱芜猪(LW)3个群体25种血液性状的染色体位点,为最后鉴定影响这些性状的因果基因提供前期基础,同时为猪抗病育种和生产提供参考。【方法】将610头杜长大三元杂猪在(180±5)日龄,336头二花脸猪和333头莱芜猪在(300±5)日龄进行统一屠宰。收集2 mL血液于抗凝管中,利用全自动生化分析仪进行25种血液性状的血常规检测。采集猪耳组织提取DNA并测浓度和质量。将质检合格的DNA样品利用Illumina 60K SNP芯片进行基因型判定。运用PLINK软件对判型结果进行质量控制,将合格的SNP标记用于后续的关联分析。使用R语言GenABEL软件包中的广义混合线性模型进行全基因组关联分析,定位影响3个群体25种血常规性状的显著性染色体位点。据全基因组关联分析结果,在Ensembl或NCBI网站上搜寻潜在的位置候选基因。【结果】杜长大猪、二花脸猪和莱芜猪三个群体通过质控的有效表型数据个体数分别为552头、325头和281头。60K SNPs经过质量控制过后,杜长大猪剩余56 216 SNPs,莱芜猪剩余49 343 SNPs,二花脸猪剩余35 974 SNPs,用于Meta分析的SNPs共有32 967。运用全基因组关联分析和Meta分析共定位到610个显著影响3个群体25种血液性状的SNPs,其中135个SNPs达基因组显著水平,475个SNPs达建议水平;分布在所有染色体上。在杜长大猪中共鉴别到32个基因组显著水平SNPs以及85个建议水平SNPs,且8种性状有基因组显著水平的SNPs,分别是淋巴细胞数目(LYM)、淋巴细胞比率(LYMR)、嗜碱性粒细胞数目(BAS)、嗜碱性粒细胞比率(BASR)、平均红细胞体积(MCV)、红细胞分布宽度变异系数(RDW_CV)、平均红细胞血红蛋白含量(MCH)和血小板分布宽度(PDW)。在二花脸猪中共鉴别到33个基因组显著水平SNPs以及139个建议水平SNPs,且9种性状有基因组显著水平的SNPs,分别是LYM、MCH、平均红细胞血红蛋白浓度(MCHC)、单核细胞数目(MON)、单核细胞比率(MONR)、平均血小板体积(MPV)、中性粒细胞比率(NEUR)、大血小板细胞(P_LCC)以及血小板压积(PCT)。在莱芜猪中共鉴别到54个基因组显著水平SNPs以及169个建议水平SNPs,且6种性状有基因组显著水平的SNPs,分别是BASR、红细胞压积(HCT)、MCH、MCHC、MCV和红细胞数目(RBC)。在Meta分析结果中,共鉴别到16个基因组显著水平SNPs以及82个建议水平SNPs,且6种性状有基因组显著水平SNPs,分别是RBC、HCT、MCH、MCHC、MCV以及MON。通过在Ensembl或NCBI网站上搜寻最强相关SNP区域内的候选基因,初步将F13A1SPTA1、DBNL、SLC25A28、CTSC基因分别确定为影响BASR、HCT、LYM、MCHC、NEUR的重要候选基因。【结论】通过全基因组关联分析和Meta分析共得到610个显著影响杜长大猪、二花脸猪和莱芜猪3个群体25种血液性状的SNP位点,初步确定F13A1、SPTA1、DBNL、SLC25A28和CTSC基因分别是BASR、HCT、LYM、MCHC和NEUR的重要位置候选基因,为解析商业猪和纯种地方猪的血液性状或免疫性疾病提供重要参考。

关键词: 猪, 血液性状, 全基因组关联分析, 候选基因

Abstract: 【Objective】 The genome-wide association of 25 hematological traits in Western DLY pigs and two Chinese Erhualian and Laiwu populations was analyzed in this study. The findings would benefit the final identification of causative genes, and provide insights into the potential molecular basis on hematological traits and provide clues for pig anti-disease breeding and production. 【Method】 A total of 610 DLY ternary hybrid pigs at (180±5) days, 336 Erhualian pigs and 333 Laiwu pigs at (300 ±5) days after birth were slaughtered at Guohong commercial abattoir. Blood samples at 2mL of each pig were immediately collected in anticoagulation tubes at slaughter. Twenty-five hematological traits were determined with commercial kits. Genomic DNA was extracted from ear tissues using a standard phenol/chloroform method, the concentration and quality were determined by NANODROP 1000 analyzer. All individuals were genotyped with Illumina porcineSNP60 Bead-Chip. After the genotype quality control by PLINK software, the GWAS were performed with the mixed linear model using GenABEL package in the R software.【Result】 After quality control of phenotype and genotype, there were 552 DLY, 325 Erhualian and 281 Laiwu left for the study, respectively; and there were 56 216 SNPs left for DLY pigs, 49 343 SNPs for Laiwu pigs, 35 974 SNPs for Erhualian pigs and 32 967 SNPs for Meta analysis. The GWAS identified 610 significant loci for 25 hematological traits, of which 135 SNPs were at the genome significant level and 475 SNPs were at the suggestive level, on all pig chromosomes in three populations. In DLY ternary hybrid pigs, 32 genome significant SNPs were identified in 8 traits (LYM,LYMR,BAS,BASR,MCV,RDW_CV,MCH and PDW) and other 85 suggestive level SNPs were also identified. In Erhualian pigs, 33 genome significant SNPs were identified in 9 traits (LYM, MCH, MCHC, MON, MONR, MPV, NEUR, P_LCC and PCT) and other 139 suggestive level SNPs were also identified. In Laiwu pigs, 54 genome significant SNPs were identified in 6 traits (BASR, HCT, MCH, MCHC, MCV and RBC) and other 169 suggestive level SNPs were also identified. In Meta results, 16 genome significant SNPs were identified in 6 traits (RBC, HCT, MCH, MCHC, MCV and MON) and other 82 suggestive level SNPs were also identified. Five promising candidate genes were found in the neighboring region of the lead SNPs at the genome-wide significant loci, such as F13A1 for BASR, SPTA1 for HCT, DBNL for LYM. SLC25A28 for MCHC and CTSC for NEUR. 【Conclusion】 A total of 610 loci surpassing the suggestive significant level were identified for 25 hematological traits on all pig chromosomes in Western DLY pigs and two Chinese Erhualian and Laiwu populations, revealing the complex genetic architecture of hematological traits. Neighboring the lead SNPs at the 54 genome-wide significant loci, five promising candidate genes were found, such as F13A1 for BASR, SPTA1 for HCT, DBNL for LYM, SLC25A28 for MCHC and CTSC for NEUR, and thus providing clues for analysis of the hematological traits and immune disease of commercial pigs and pure indigenous breeds.

Key words: pig, hematological traits, genome-wide association study, candidate gene