转录因子MYB调控采后低温李果肉红变的机制研究

Plum is one of the most favourable fresh fruits, with colorful appearance and multiple nutrients. However, abnormal flesh reddening (yellow as normal) usually occurs in those plums with black, purple or red skin during storage at low temperature and the subsequent removal to shelf-life, which strongly caused considerable losses of the fruit in the cold-chain transportation and for marketing supply. Thus, in this project, the transcriptomic analysis is to be performed to identify differences of the gene expression related to the anthocyanin biosynthesis pathway, and the gene expression of the main transcription factors of plums responding to low-temperature storage. The correlation among these gene expressions in the flesh reddening of plum fruit is also to be elucidated. The MYB transcription factors that respond to the low-temperature stress as well as can regulate the biosynthesis of anthocyanins are to be selected and identified for further study. MYB gene cloning and the expression analysis are to be conducted. The recognition and binding of the key MYB transcription factor and the specific nucleotide region of the promoters of downstream anthocyanin synthesis target genes, and the transcriptional activation properties of MYB are to be explored. The function of the MYB transcription factor binding with its interaction protein and their act on the regulation network on the anthocyanin synthesis are to be investigated too. Thereby, the mechanism of MYB transcription factor on regulating the low-temperature induced flesh reddening in plum fruit after harvest is to be clarified. Based on these studies, the effects of postharvest treatments on the gene expression and regulation function of the key MYB transcription factor in plum fruit stored at low temperature are to be explored. The expected results from this project would provide a novel theoretic base for controlling of postharvest quality deterioration of plum fruit during storage at low temperature, as well as provide a reference for postharvest regulation of the anthocyanin biosynthesis, the active and functional components in the plum fruit.

李果实色泽艳丽、营养丰富，是优良的鲜食水果种类之一。但是，黑/紫/红皮等类型李在采后低温贮藏及贮后常温货架期极易出现果肉红变现象（正常果肉为黄色），严重影响李的贮藏、冷链流通和市场供应。为此，本项目拟通过转录组测序，研究低温李果实花色苷合成途径基因、主要转录因子基因的表达差异，以及它们与采后低温李果肉红变的关联响应关系，确定能够应答采后低温激发诱导、调控花色苷合成的相关MYB转录因子。进行MYB基因克隆和表达特性分析，研究关键MYB转录因子与下游花色苷合成靶基因启动子区域特异核苷酸的识别与结合，以及转录激活特性；研究MYB与互作蛋白的作用及其对花色苷合成的网络调控，从而阐明MYB转录因子调控采后低温激发诱导的李果肉红变的机制。基于此，探索采后处理对关键MYB转录因子基因表达及其调控功能的影响，为李果实低温贮藏品质的控制提供新理论依据，为李果实活性功能成分花色苷代谢的采后调控提供参考。

不同类型李果实低温贮藏特性差异较大，黑/紫/红等深色果皮类型李品种如‘黑宝石’、‘澳洲14’、‘安哥诺’等，在常温贮藏时果肉逐渐变为琥珀色，但在低温贮藏及解除低温后的货架期（常温）极易出现急剧、快速的果肉红变现象，这种采后低温对果实性状的调控尚未得到研究。本项目研究了果肉红变的发生机制，并探索有效的方法对贮藏红变现象进行调控以保持果实的品质。液谱-质谱分析发现，红变物质组分为大量的矢车菊素3-O-葡萄糖苷和极少量的矢车菊素3-O-芸香糖苷。代谢组学分析表明，低温贮藏刺激苯丙烷代谢和花色苷代谢产物积累。生化分析表明，低温贮藏激活了花色苷代谢相关酶活性。伴随着花色苷的积累，李果实的乙烯释放速率出现迸发。转录组分析表明，低温贮藏刺激冷信号、乙烯合成与信号转导、主要转录因子基因，花色苷生物合成途径中结构基因的显著上调表达。利用瞬时过表达和基因沉默方法研究发现，乙烯响应因子ERF1B参与了转录因子MYB10.1和bHLH3对李果肉红变的调控作用，可能介导了李果实对低温刺激的响应。通过分子生物学方法研究，明确了MYB与互作蛋白的作用，为深入理解采后低温调控李果实性状的转录网络调控机制提供了依据。本项目进一步研究了采后1-MCP熏蒸、PVC包装、褪黑素处理对低温诱导‘Friar’李果肉红变的调控作用及对转录表达的影响，为调控采后果肉色泽性状提供了技术参考。研究结果为我国李果实采后贮藏品质控制与花色性状调控提供了重要依据。项目执行过程中发表8篇研究论文，培养了多名研究生，对李果实农户进行了技术指导和实践应用。