虾夷扇贝性别分化关键基因的鉴定、染色体定位及功能分析

Patinopecten yessoensis is an important aquaculture scallop species in the north of China, which contribute for the scallop culture structural adjustment and development. In recent years, there is consensus that hermaphrodite could be used for variety improvement by constructing inbred lines, but we are only beginning to grasp the research on the characteristic of shellfish sex differentiation. In this.study, by using the scallop Patinopecten yessoensis as the research model with important breeding status, hermaphrodite phenomena and sound basis of research, we aim to investigate the genetic mechanism of hermaphrodite formation on the transcriptomic level. The research plan includes: First, by using the high-throughput RNA-seq and WGCNA method of scallop gonad and larvae of different early development stages, screening and identification of genes correlated with sex differentiation; Second, by using RT-PCR and in situ hybridization, key genes about sex differentiation and their temporal and spatial expression model, and alternative splicing sites; Third, by using bioinformatics, construction of key genes co-expression network; Forth, by using FISH technique, key genes chromosomal location; Final, by using gene silencing technique and model organisms with gene editing, gene function in sex differentiation. This study will not only lay an important theoretic foundation for further investigations on shellfish sex differentiation, but also demonstrate that hermaphrodite is an excellent research model for studying shellfish sex differentiation. Our study would provide necessary technical and theoretical support for Patinopecten yessoensis further genetic improvement and sex differentiation regulatory pathway construction.

虾夷扇贝是我国北方扇贝养殖业结构调整及辅助发展的重要养殖品种。近些年来，利用养殖贝类中雌雄同体构建自交家系改良品种已在科研人员中达成共识，但有关贝类性别分化的研究才刚开始。本项目选取养殖地位重要、有雌雄同体现象、研究基础好的虾夷扇贝作为性别分化的研究模型，从转录组水平上研究雌雄同体形成的遗传机制。主要内容包括：利用高通量转录组测序技术和WGCNA方法，筛选鉴定性别分化相关基因；利用RT-PCR和原位杂交技术构建基因时序表达模式和转录本结构示意图，确定性别分化关键基因；利用生物信息学分析方法构建基因共表网络图；利用FISH技术进行基因染色体定位；利用基因沉默技术和基因编辑后的模式生物验证关键基因在性别分化中的调控作用。本项目的研究成果将为海洋贝类性别分化的理论研究提供基础资料和新的模式生物，为虾夷扇贝遗传改良及性别分化调控通路图的构建提供理论和技术支持。

虾夷扇贝是我国北方扇贝养殖业结构调整及辅助发展的重要养殖品种。为研究贝类性别分化的分子机制，本项目对养殖地位重要、有雌雄同体现象、研究基础好的虾夷扇贝作为性别分化的研究模型，从转录组水平上研究雌雄同体形成的遗传机制。主要内容包括：利用高通量转录组测序技术和基因共表达网络分析方法（WGCNA），筛选鉴定性别分化相关基因49个；构建了虾夷扇贝性别分化遗传通路图，其中Dmrt 1基因的可变剪接是性别分化方向的转换开关，决定性别分化方向；利用荧光定量PCR和原位杂交技术构建性别分化关键基因Dmrt1 和Foxl 2 的时序表达模式，Dmrt 1 的表达方式调控了Foxl 2 的表达；雌雄染色体核型分析结果表明，不同于栉江珧，虾夷扇贝尚未出现性染色体的初级分化；生殖腺成熟早期RNAi干扰的初步结果表明，Dmrt1 基因对生殖腺RNAi干扰及其表达蛋白生殖腺注射实验正在开展，已获得一些初步的结果表明Dmrt1基因确实对虾夷扇贝性别分化有调控作用。本项目的研究成果将为海洋贝类性别分化的理论研究提供基础资料和新的模式生物，为虾夷扇贝遗传改良及性别分化调控通路图的构建提供理论和技术支持。