综合组学策略鉴定猪骨骼肌肌纤维类型构成相关基因及其适应性进化机制

Pig has been under artificial selection for long time, and it is a domestic animal with clear evolution and domestication history compared to others. Pig is one of important livestock for meat production. Muscle fibers (I type and II type) compose skeletal muscle and their number and diameter determine the meat production capability. I and II type fiber composition has been changed greatly due to captivity and high density selection of long time, and it is shown between wild, domestic and commercial developed pig breeds. Based on the previous research on genome wide selection sweep, this study will choose Tongcheng, Yichang Wild pigs and Large White to investigate the molecular mechanism on muscle fiber type composition for selection adaption from both genome and transcriptome level. Expression profiling of soleus and gastrocnemius muscle tissues from the studied pigs will be analyzed using RNA-seq technique, and then transcriptome difference are clarified. More SNPs will be increased within candidate genome region and selection sweep analysis will be implemented for fine mapping. Furthermore, allelic expression imbalance and gene function analysis approaches will be executed for biological function verification of candidate genes. The selection evolution pattern, molecular characterization and mechanisms of candidate gene and pathways will be demonstrated by means of population genetics and evolution genome statistical methods. The findings will be expect to help reveal the complex phenotypic evolution in pigs and contribute to better understanding of adaptability formation under artificial selection.

猪是经长期人工选择、且进化及驯化历程较为清晰的肉用家养动物，高产肉性能是其重要机能发生改变的表征之一。肌纤维（包括红肌和白肌纤维）是骨骼肌基本组成单位，其数量和直径决定着产肉能力高低。长期圈养及对产肉性能的高强度选择使得红肌/白肌纤维类型构成发生显著改变，在野猪、地方猪和培育猪之间呈现出巨大差异。本研究拟在前期成果基础上，选取我国地方通城猪，以宜昌野猪和瘦肉型大白猪为对照，从基因组和转录组水平对猪肌纤维类型构成变异的适应性分子机理进行探讨。通过RNA-seq技术对三个猪种的比目鱼肌（以红肌为主）和腓肠肌（以白肌为主）组织表达检测分析，揭示转录组比较差异；增补候选基因组区域SNP 密度，通过基因组选择清扫分析以精细确定候选基因；利用等位基因表达不平衡和基因功能研究技术，初步阐明候选基因生物学功能；综合群体遗传学和进化基因组学统计分析手段，探讨相关基因及通路的选择进化模式、分子作用规律及机制。

高强度肌肉生长是人工选择条件下猪适应性进化身体机能发生改变的重要因素，肌纤维数目与组织学特性是决定瘦肉产量与肌肉品质重要理化基础，因此本研究拟通过多组学手段鉴定与猪肌纤维以及肉质性状相关的功能基因及致因突变。我们通过对野猪、通城猪、陆川猪、大白猪和杜洛克猪等不同品种猪的选择性清扫分析在家猪基因组上鉴定到25个选择信号区域，其中23个位于X染色体；通过多品种全基因组SSR多态性分析发现我国地方猪群体内杂合度与多态信息含量整体上比瘦肉型猪高，说明地方猪品种内选育提高空间较大。通过大规模屠宰测定，我们开展了较大群体肌肉组织的RNA-seq测序和eQTL分析鉴定到15个与肌纤维性状显著相关且受cis-eQTL调控的候选基因，其中PHKG1基因内含子剪接位点突变与屠宰后24小时肌肉pH值和肉色显著关联；分别检测到影响肌纤维特异表达的MYLPF、TNNC2和TNNI1基因的cis-eQTL，初步鉴定了各自的功能突变，这些功能突变的多态性与肌纤维及肉质性状存在不同程度的相关。项目合作单位研究了GPAT3和ssc-miR-7134-3p两个与生长性状密切相关的功能基因，发现GPAT3基因启动区一个SNP改变了其启动子活性与表达水平；而ssc-miR-7134-3p可以降低MAPK4基因的表达。因此，本研究结果为探讨猪肌肉生长及肌纤维特性的适应性进化分子机制提供了重要参考，同时，本研究鉴定到的15个受cis-eQTL调控的候选基因为肉质性状遗传改良提供了基因素材，已经鉴定到的几个功能突变将有望应用于猪肉质性状的遗传改良。