大黄鱼(Larimichthys crocea)新品种“东海1号”体长相关的DArT标记筛选

IDENTIFICATION OF BODY-LENGTH-RELATED DART MARKERS IN NEW CALTIVAR “DONGHAI NO.1” OF LARGE YELLOW CROAKER LARIMICHTHYS CROCEA

大黄鱼(Larimichthys crocea)是我国重要的海产经济鱼类。因多年未加选育的累代养殖,养殖大黄鱼生长缓慢,性早熟、免疫力低下,亟需对养殖大黄鱼进行遗传改良。多样性芯片技术(Diversity arrays technology,DAr T)具有高通量和低成本的显著特点,不需要明确物种的基因组DNA序列信息,因而广泛应用于动植物遗传图谱制作和遗传多样性分析。本研究旨在采用DAr T技术鉴定与"东海1号"大黄鱼体长相关分子标记。首先随机测得"东海1号"大黄鱼199个个体体长,均值为13.45cm,符合正态分布(P>0.05),"极端大群体"和"极端小群体"之间差异显著(P<0.01)。然后采用7组限制性内切酶组合(Pst I/Alu I、Pst I/Ban II、Pst I/Bsp1286I、Pst I/Bst NI、Pst I/Hae III、Pst I/Rsa I、Pst I/Taq I)分别进行酶切,获得基因组代表性DNA片段并用于文库构建。根据多态性克隆数和多态性率确定Pst I/Rsa I酶切组合为最优降低基因组复杂度方法。之后从Pst I/Rsa I基因组代表性DNA片段文库中扩增获得3360个片段,以此为多样性芯片探针进行芯片点制,杂交筛选获得18个大黄鱼体长相关DAr T候选标记。经过新群体样本再次验证,仍有8个DAr T标记与体长相关。本研究有助于选育生长性状优良的大黄鱼群体。

Large yellow croaker（Larimichthys crocea） is an important marine economic fish in China. However, in recent years, many commercial traits have been degrading due to fast development of fish farming, resulting in lower growth rate, earlier sex maturation, and lower viability. Thus, the need is urgent in genetic improvement of farmed large yellow croaker. Diversity arrays technology（DAr T） provides a high-throughput and low-cost whole-genome genotyping platform for the detection and scoring of hundreds of polymorphic loci without any need for prior sequence information. DAr T is widely used to create genetic maps and analyze genetic diversity of animals and plants. We used DAr T to identify body-length-related molecular marks of the Donghai No.1 caltivar of large yellow croaker. The body lengths of 199 fish were randomly measured and the average was 13.45 cm in conformation to normal distribution. The maximal and minimal length groups were significantly different（P〈0.01）. Pst I representations were digested with seven pairs of different frequent cutters（Pst I/Alu I, Pst I/Ban II, Pst I/Bsp1286 I, Pst I/Bst NI, Pst I/Hae III, Pst I/Rsa I, Pst I/Taq I） and used for libraries construction. Pst I/Rsa I was selected as optimal complexity reduction method according to the numbers of unique clones and polymorphism levels. An amount of 3360 Pst I representations as probes was amplified from the Pst I/Rsa I library for array preparation. Eighteen DAr T candidate marks were screened by hybridization and data statistics. The new selected population was used to verify the candidate marks, and eight DAr T markers were significantly associated with body length. These basic data highlight potential application of DAr T for marker-assisted selection of large yellow croaker with good growth traits.