大额牛独特肉质性状形成的分子调控网络及cSNP功能位点分析

大额牛（Bos frontalis）为半家养的独立牛种，具有典型的肉用性能。前期研究发现大额牛具有极细的肌纤维和极低的肌内脂肪含量等独特肉质性状。本研究选择云南本地黄牛为对照，应用RNA-Seq检测两品种间第260天胎儿和屠宰日龄牛肌肉组织mRNA序列，高通量筛选两种群肌肉组织差异表达基因、新基因、编码SNP(cSNP)和可变剪接体；通过贝叶斯法构建基因网络，对关键基因进行荧光定量PCR和western blot表达量验证；克隆关键差异表达基因和重要cSNP所在候选基因全基因序列；在两品种各30个个体中初筛SNP，对SNP进行单倍型分析，确定标签SNP（TagSNP），通过MassArray进行大样本TagSNP分型，确定优势SNP；开展编码SNP和调控SNP功能预测，并通过凝胶阻滞（EMSA）验证调控位点功能，发掘肉质功能基因和位点。研究结果为大额牛种质特色基因的开发和利用奠定了基础。

The gayal, as independent Bos specie, distributes only in Yunnan in China, likes eating bamboo, still keeps semi-domestication and possesses typical meat physique. Our previous work revealed unique meat quality traits of meat quality such as very fine muscle fiber and very low intramuscular fat content. In this study, we plan to select Yunnan cattle as contrast control, determine mRNA sequences of muscle of day 260 fetus with strongest cell differentiation and adult individuals with a large fatty deposition using RNA-seq to screen differential expression gene, novel gene, cSNP and alternative splicing transcripts; construct Bayes gene network and validate expression level of important genes using QPCR and western blot; clone key differential expression gene and candidate gene containing significant cSNP, prescreen SNP in 30 individuals in respective group, analyze haplotyes to identify TagSNP, and type TagSNP using MassArray with a large sample to identify dominant SNP; predict function of cSNP and non-missense SNP, and validate function of non-missense site using EMSA, and consequently find functional genes and sites. These results will settle the basis for exploring and utilizing characteristic gene of the gayal.