野生西瓜钾高效吸收相关基因的发掘及功能验证

Potassium is an essential macroelement for plant growth and development. The limited potassium in the soil and the shortage of potassium fertilizer are important limited factors for the sustainable agriculture in China. Watermelon is an important horticultural crop, and the need of potassium for plant growth and development of watermelon is high. Our previous study found that the potassium uptake capacity and low potassium tolerance of wild watermelon is generally higher than that of cultivated ones, and the low potassium tolerance of cultivated watermelon can be enhanced by grafting onto wild watermelon rootstocks. In the present project, the wild watermelon cultivar'Yongshi' with high potassium uptake capacity and the cultivated watermelon cultivar'8424'with low potassium uptake capacity were used as materials. The candidate genes responsible for the high potassium uptake capacity of wild watermelon will be identified, by studying the low potassium responsive gene expression profiling of root using the high-throughput sequencing technique, in combination with the bioinformatics analysis. The spatial and temporal expression of candidate gene will be conducted by Real-time PCR. The full-length candidate gene will be cloned. On this basis, the functional verfication of candidate genes will be conducted through the genetic transformation of Arabidopsis. It is expected that the implementation of this project will obtain the key genes for high potassium uptake capacity of wild watermelon, and lay a foundation for the elucidation of molecular mechanism for the low potassium tolerance of wild watermelon. Meanwhile, the result will also provide the theoretical basis for the enhanced low potassium tolerance of cultivated watermelon by grafting onto wild watermelon.

钾是植物生长发育所必需的大量元素。土壤钾不足和钾肥资源贫乏是我国农业生产可持续发展的重要限制因子。西瓜是一种重要的园艺作物，对钾素需求量大。项目组前期研究发现，野生西瓜对钾的吸收能力和耐低钾性能普遍强于栽培西瓜，利用野生西瓜为砧木嫁接可以显著提高栽培西瓜的耐低钾能力。本项目以前期筛选出的钾高效吸收型野生西瓜'勇士'和钾低效吸收型栽培西瓜'8424'为材料，利用高通量测序技术研究低钾胁迫下根系基因表达谱的变化，进行生物信息学分析，确定野生西瓜钾素高效吸收候选基因。利用Real- time PCR技术研究候选基因时空表达模式；克隆候选基因全长；通过转化模式植物拟南芥的方法对重要候选基因进行功能验证。本项目的实施可望获得野生西瓜钾高效吸收关键基因，为揭示野生西瓜耐低钾的分子机制奠定基础，同时也为野生西瓜嫁接提高栽培西瓜的耐低钾性能提供理论依据。

钾是植物生长发育必需的大量元素。土壤钾不足和钾肥资源贫乏是我国农业生产可持续发展的重要限制因子。西瓜是一种重要的园艺作物，对钾素需求量大。本项目以前期筛选出的钾高效吸收型野生西瓜‘勇士’和钾低效吸收型栽培西瓜‘早佳8424’为材料，利用RNA-Seq和QPCR技术研究了低钾（0 mM K+）早期（6 h）和后期（120 h）根系基因表达谱的变化，结合传统生理手段测定了光合作用、矿质营养、抗氧化和内源激素等参数，提出了野生西瓜‘勇士’适应低钾胁迫的分子生理模型：根系发达，K+转运蛋白基因表达丰度高，对根际低浓度K+的亲和能力好，且能有效利用Mg2+和Ca2+代替K+；抗氧化能力强，生长相关的激素(IAA、GA3)与衰老相关的激素(ABA、ETH)之间的平衡维持较好；野生西瓜‘勇士’能够在非逆境下高量表达很多抗逆相关的基因，而在长期低钾胁迫下能够下调表达与胁迫响应和防御反应相关的基因，从而使更多的能量被根系生长所利用，而强大的根系可以有效地吸收K+，从而逐渐适应低钾胁迫。基于RNA-Seq生物信息学分析和前人钾营养研究进展，筛选出野生西瓜钾高效吸收候选基因30个。克隆了野生西瓜钾高效吸收候选基因Cla012760（HAK5）的全长，长度为2400bp，将此基因转到模式植物拟南芥hak5突变体中，已经获得了T2代植株，目前正在进行耐低钾性鉴定，评价野生西瓜Cla012760基因在钾高效吸收中的功能。本项目的实施部分解析了野生西瓜耐低钾的分子机制，获得了钾高效吸收的30个候选基因，为进一步揭示野生西瓜耐低钾的分子机制奠定了基础。本项目已经发表论文4篇，其中SCI论文2篇，培养毕业博士和硕士研究生各1名，获得湖北省科技进步二等奖和三等奖各1项。项目主持人作为大会秘书协助会议召集人成功组织召开了“第一届国际园艺学会蔬菜嫁接研讨会”（2014.3.17-21）。