长牡蛎黑色素合成通路关键基因鉴定及其影响壳色多态性的分子机制

长牡蛎（Crassostrea gigas）是世界上养殖范围最广、产量最大的海洋贝类，在我国海产贝类养殖业中占有十分重要的地位。本项目拟选用经过6代定向选育、性状遗传稳定的4种壳色（白色、黑色、金黄色和紫色）长牡蛎为研究素材，通过高效液相色谱分析，查明长牡蛎外套膜和贝壳中黑色素及其前体的种类及含量比例；通过高通量转录组分析，筛查长牡蛎黑色素合成通路关键基因及其非编码RNA（lncRNA和microRNA）；利用RACE技术，克隆黑色素合成通路关键基因，并通过qPCR，原位杂交和免疫组化方法，分析其表达特性；采用RNA干扰技术对关键基因及其非编码RNA进行功能验证及调控分析，解析关键基因在黑色素合成通路中的功能及其表达调控网络；通过黑色素合成通路关键基因遗传变异与壳色多态性的关联分析，解析壳色多态性产生的分子基础。研究结果将为揭示长牡蛎壳色形成的分子机制、培育壳色相关优良品种奠定坚实基础。

As a commercially important marine bivalves, the Pacific oyster (Crassostrea gigas) is a major global aquaculture species, ranking first in the production among all aquaculture shellfish species. In a breeding program, four lines of C. gigas with inherited steadily white, black, golden, and purple shell colors have been established through successive selection for six generations. In this study, the melanin type and precursor in shells and mantles will be identified and quantified using high-performance liquid chromatography analysis. To identify the gene and non-coding RNA (lncRNA and microRNA) mainly involved in melanin biosynthesis pathway, the transcriptome in four shell color lines will be sequenced. The genes which may play key roles in melanin biosynthesis pathway will be cloned and characterized by RACE, qPCR, in-situ hybridization, and immunohistochemical staining. RNA interference will be performed to analyze the expression and regulatory network of the genes and non-coding RNA ((lncRNA and microRNA) involved in melanin biosynthesis pathway. Through analyzing association between genetic variation in the key genes and shell color polymorphisms, the molecular base of shell coloration will be clarified. This work will benefit comprehension of the molecular mechanism of shell color variation, and selective breeding in terms of shell color traits in C. gigas.