大白菜平塌突变体fg1关键基因的功能鉴定及调控机理

Chinese cabbage (Brassica rapa L.ssp. pekinensis) is an important vegetable crop and its leafy head is the main edible organ. However, the molecular mechanism of leafy head development is unclear. Based on one ethylmethane sulfonate (EMS)-induced flat growth mutant (flat and divergent leaf type) with the same genetic background as Chinese cabbage ‘A03’, the heredity analysis showed the mutant character was controlled by a pair of recessive gene and high-throughput sequencing (MutMap) preliminarily mapped mutant loci. The single plant in segregation population with extreme traits will be sequenced to confirm the mutant gene locus and sequence features which were detected in MutMap. RNA-seq will be used to identify the regulatory network of differentially expressed genes and spatio-temporal patterns of those genes in heading biological process of Chinese cabbage. The leafy head morphology, plant hormones and leaf anatomical structure will be analyzed to explore the signaling pathway during leaves heading. The function of the key gene will be confirmed via the CRISPR gene-editing technique together with vernalization-infiltration methods. This study will not only identify regulatory genes of leafy head, but also clarify the function of the new key gene and the signaling pathway of leaves heading, which will reveal the regulation mechanism of the leafy head formation. This project will facilitate the in-depth research of the molecular mechanism responsible for leafy head development and provide a technological base for molecular marker assisted breeding of the heading trait in Chinese cabbage.

大白菜是以叶球为主要食用器官的重要蔬菜作物，但其叶球发育的分子调控机理尚不明确。本项目以申请者实验室创建的具有相同遗传背景的大白菜平塌（叶片平塌、散叶型）突变体fg1及其野生型为材料，在已确定该突变性状由一对隐性基因控制，并初步定位了突变位点的基础上，进一步通过分离群体极端性状单株测序验证突变基因位点，确定其序列变异特点；利用RNA-seq解析大白菜结球相关基因的表达调控网络和时空变化规律；同时分析叶球发育、内源激素和叶片解剖结构的表型差异，初步明确大白菜结球的信号调控途径；结合CRISPR基因编辑和真空渗透转化体系，开展结球关键基因的功能验证等研究。该研究不仅对叶球发育调控基因进行鉴定，而且对新发现关键基因的功能及调控结球的信号途径解析研究，揭示关键基因对大白菜结球的调控机理。项目的开展有助于推进大白菜叶球发育的分子机制的深入研究，也可为大白菜结球性状分子标记辅助选择育种提供技术基础。

大白菜（Brassica rapa L. ssp. pekinensis）为十字花科芸薹属植物，主要以叶片作为食用器官，变异类型丰富，从最初的散叶型，逐渐进化为半结球型、花心型和结球型。大白菜包心期，球叶的叶夹角变小，叶片边缘向内卷曲，形成叶球。叶球发育过程受许多因素影响，是一个极为复杂的生物学过程。.本研究以甲基磺酸乙酯（EMS）诱变构建的大白菜突变体库中筛选出的平塌突变体fg-1为试材，在鉴定该突变体农艺性状以及叶片激素含量的基础上，取突变体fg-1及其野生型（Wild type，WT）大白菜包心叶片4个部位，借助转录组测序（RNA-Seq）技术，在基因转录水平揭示突变性状的表达调控机制；通过构建F2和BC1分离群体，分析突变基因的遗传特性；利用MutMap技术定位候选基因，并对该候选基因的结构特点及其表达调控进行分析，进而揭示fg-1叶球平塌的分子调控机制。本研究对进一步挖掘大白菜叶球相关新型基因，实现蔬菜株型定向育种提供重要理论依据。