文蛤生长的关键基因及其表达调控机理研究

The hard clam (Meretrix meretrix) is a commercially important maricultured shellfish in China. Growth is one of the most important goal traits in the clam selective breeding. Discovery and identification of key growth factor genes are the primary prerequisite for in-depth study on the molecular mechanism of growth regulation and the cultivation of high-yield varieties. A large number of studies on other animals have found that a series of cytokines, including MSTN,BMPs,IGF1, EGF and etc., regulating the growth of muscle, bone and epithelial, were the main factors affecting growth trait. This study, based on the transcriptome library of Meretrix meretrix, intend to clone the full-length cDNA, intron and promoter sequences and to analyze the structural features and genetic variations of growth candidate genes, like MSTN, BMPs, IGF1, EGF and EGFR, using RACE and Genome Walking techiques. By qRT-PCR method, we analyze the gene expression patterns in different tissues and at different developmental stages to confirm the close relationship between the genes and growth traits. Using the rapid growth families and groups as experimental material, by large-scale sequencing and HRM technology, the growth-related SNP markers will be screened and identified to build G-P model between genotype and phenotype. The purpose of the study is to explore the molecular mechanism of growth differences and to provide theoretical basis and technical support for molecular marker-assisted selection and molecular design breeding of the hard clam.

文蛤是我国沿海滩涂养殖的重要经济贝类，其生长是育种研究和实践首要关注的目标性状。发掘文蛤生长的关键基因，是深入研究生长调控分子机制和开展高产良种培育的重要前提。对多种动物研究发现，MSTN、BMPs、IGF1、EGF等是调控肌肉、骨骼、上皮生长的重要细胞因子，也是影响生长性状的重要因素。本项目拟以文蛤转录组为基础，利用RACE和Genome-Walking技术克隆MSTN、BMPs、IGF1、EGF和EGFR等多个生长候选基因的cDNA全长、内含子和启动子，分析基因的结构特征和遗传变异；采用qRT-PCR分析其在不同组织、不同发育阶段的时空表达，证实基因与生长的密切关系；以快速生长家系和群体为试材，利用大规模测序和HRM技术筛查和鉴定生长的关键基因和SNP位点，构建基因-表型相关的G-P模型，为SNP用于文蛤分子标记辅助育种和分子设计育种提供理论指导。

文蛤是我国沿海滩涂养殖的重要经济贝类，而生长是育种中首要关注的目标性状，发掘和鉴定生长关键基因是深入研究生长调控分子机制和大力开展高产良种培育的重要前提。MSTN、IGF、EGF、FGF等是调控肌肉、上皮、纤维细胞生长的重要细胞因子，是影响生长性状的重要因素，利用海量基因组、转录组信息发掘生长相关基因并阐明其表达调控机制，已成为分子遗传育种的重要研究方向和内容。本项目以Roche 454和Illumina Solexa测序的文蛤转录组为基础，利用注释基因信息筛选生长候选基因EST，构建生长候选基因信息库；利用RACE和Genome Walking技术克隆MSTN、BMPs、Smads、HDACs、IGF等生长候选基因的cDNA全长、内含子和启动子，分析其结构特征和系统演化规律；采用qRT-PCR分析其在不同组织、不同发育阶段的表达规律，证实基因与生长的关系；利用大规模测序和基因分型技术筛选生长相关SNP标记，为文蛤分子标记辅助育种提供理论依据。 .项目实施四年来，取得了一系列重要成果，（1）系统地对文蛤MSTN、BMPs、Smads、HDACs等9个生长候选基因进行了分子克隆、基因结构解析；（2）分析了9个候选基因在文蛤不同组织器官、不同发育阶段及不同胁迫刺激中的时空表达规律；（3）筛查到文蛤生长关键调控基因5个、生长紧密相关SNP标记 11个，并开发转录组源SNP标记222个；（4）利用群体选育结合分子辅助育种技术，培育出文蛤“万里红”、“万里2号”2个快速生长新品种，养殖生产性能测试优良性状表现明显。相关研究成果已在国内外学术期刊上发表或接受研究论文8篇，其中SCI论文3篇，出版专著1部；申请并授权发明专利1项，培养硕士研究生5名，获国家审定文蛤新品种2个；获省部级二等奖2项和市级一等奖1项。研究结果可为文蛤生长性状的分子调控机理研究和分子标记辅助育种实践奠定重要基础。