大白菜干烧心病发生中的盐钙互作及钙依赖蛋白激酶(CDPKs)调控机制研究

Tipburn is the main reason affecting the internal quality of Chinese cabbage in the anniversary production and marketability of Chinese cabbage. Physiological calcium deficiency is the root cause in the occurrence of tipburn. Calcium-dependent protein kinase (CDPKs) plays a key role in the process of calcium absorption and transport. Tipburn disease has a quantitative trait controlled by multiple genes, and largely impacted by the cultural environment. The study between the relationship of high salt and tipburn occurence of Chinese cabbage, the molecular mechanism of sodium-calcium interaction, the contents of calcium-dependent protein kinase, the contents of calcium-binding protein and anti-tipburn Chinese cabbage have rarely been reported. It was studied on the activity of calcium-dependent protein kinase and the gene expression difference between resistant and susceptible materials under the condition of salt and calcium stress. The intrinsical links were analyzed among the contents of calcium-dependent protein kinase, the contents of calcium-binding protein and the changes of sodium-calcium absorption and distribution. The CDPK family genes of Chinese cabbage were identified, to quantitative analysis of its protein, and ascertained the differences in the level of the expression and translation of gene, and which revealed the intrinsic regulatory mechanism of the anti-tipburn variety. It provided reference for modern molecular breeding of anti-tipturn of Chinese cabbage.

大白菜干烧心病表现球叶隔层坏死或干枯，极大影响其商品性和产量，已成为大白菜周年生产供应中的一大主要病害。生理性缺钙是导致干烧心病发生的根本原因。钙依赖蛋白激酶类蛋白在钙素吸收转运中起着关键作用。干烧心病是多基因控制的数量性状，受栽培环境影响大。针对高盐与大白菜干烧心发生的关系、钠-钙间互作的分子机制、钙依赖蛋白激酶和钙结合蛋白含量与大白菜干烧心病的内在联系和关联分析鲜有报道。本研究以多年筛选出的大白菜抗、感干烧心品系为试材，研究盐胁迫、钙胁迫条件下，抗、感材料在干烧心病发生过程中钙依赖蛋白激酶基因表达的差异，分析钙依赖蛋白激酶基因与钠-钙吸收分布变化的内在联系，对大白菜CDPK家族基因进行鉴定，并通过蛋白定量分析，探明其在翻译水平的基因表达差异，近而揭示其对抗干烧心病能力的内在调控机制，为大白菜抗干烧心病的现代分子育种提供参考。

本项目以多年筛选出的大白菜(Brassica campestris L.ssp pekinensis)抗、感干烧心品系为试材，研究盐胁迫、钙胁迫条件下，抗、感材料在干烧心病发生过程中的生长状况和元素吸收分配差异，研究结果表明了抗病品系元素向叶片运输能力强于感病品系，且叶柄和叶片中钙含量与干烧心病发生显著相关，感病品系叶片和叶柄中积累的钠元素影响了钙素的吸收，从而造成感病品系叶片中钙元素吸收障碍而过早发生干烧心病；本项目研究完善了低钙营养液胁迫下大白菜干烧心病水培鉴定体系，以大白菜高抗干烧心病DH系C8、高感干烧心病DH系651A为试材，对体系重要步骤和参数进行了优化，该鉴定体系解决了长期低钙培养下幼苗黄化和症状不典型问题。以水培鉴定体系和大白菜抗、感材料在干烧心发病过程中症状表现为基础，利用转录组测序分析抗、感材料在发病过程中的差异表达基因，并对CDPK基因进行鉴定，共发掘新基因1,183个，其中972个得到功能注释。基于比对结果，进行基因表达量分析，结果表明，感病品系处理9天时与对照相比共有差异表达基因736个，其中上调表达361个，下调表达375个；抗病品系处理9天时与对照相比共有差异表达基因819个，其中上调表达247个，下调表达572个；处理期0、3、6、9天时，感病处理组与对照组共有的差异表达基因为22个；抗、感病处理组与对照组共有的差异表达基因为12个；挖掘CDPK相关的差异表达基因53个，其中6个通过参与非生物胁迫信号途径调控基因表达。本项目研究明确了大白菜抗、感品系在干烧心发病过程中的盐、钙间互作关系，抗病品系和感病品系在元素向叶片运输能力差异指标具有重要的育种实践价值；挖掘了大白菜抗、感材料在干烧心发病过程中差异表达基因，鉴定了CDPK基因及其在干烧心发病过程中的表达差异，为大白菜抗干烧心病的现代分子育种提供参考。