葡萄一年两收栽培二次果成花的生理分子机制及调控研究

The two-crop-a-year viticulture makes optimum use of the abundant light and heat resources in North China. It overcomes the difficulty of insufficient chilling requirement. The first and the second fruit maturity stage of the two cropping was during the earliest and the latest stages of grape maturity period in the whole country, respectively, which perfectly fills the cap of market requirements and greatly improves the economic benefits. By now the two-crop-a-year viticulture has been spread across several provinces in North China such as Guangxi, Fujian, Yunnan, Hainan, Guangdong, etc. However, unstable inflorescence differentiation is the main bottleneck problem that threatens the production benefits and further application of the two-crop-a-year viticulture. A deeper understanding of inflorescence differentiation and its regulatory factors to figure out how to improve inflorescence fertility is vital to stable yields in two-crop-a-year viticulture. 'Summer Black' grapes are selected as materials in this study. The morphological inflorescence differentiation process and level changes of endogenous hormones are studied. Besides, genes regulating grapevine flowing are cloned and detected at the transcriptional level to figure out the temporal and spatial expression patterns. Meanwhile, several plant-growing regulators are prayed and different degrees of summer pruning are applied as different treatments. The morphological inflorescence differentiation in the second grapevine crop as well as the physiological and molecular mechanisms response to different treatments is studied. Based on the above studies, we aim to find suitable plant-growing regulators praying and proper degree of summer pruning. The results will provide important theoretical basis for maintaining stable yields in grape two-crop-a-year cultivation.

葡萄一年两收栽培模式充分利用了南方丰富的光热资源，克服了一收模式下葡萄需冷量不足的难题。两收葡萄的成熟期分别处于全国葡萄成熟的最早和最晚时期，弥补了市场空缺，大大提高了经济效益。目前已在广西、福建、云南、海南、广东等地推广。但二次果成花不够稳定、冬果产量难以保证是两收栽培技术进一步发展的瓶颈。深入了解葡萄花芽分化规律和影响二次果花芽分化的主要因子，找到促进葡萄二次果花芽分化的措施是葡萄一年两收稳产的关键。本研究拟以'夏黑'葡萄为试材，通过形态解剖研究新梢二次果花芽分化进程，同时测定新梢冬芽激素含量变化，克隆成花关键基因并研究其时空表达特征及水平，同时，研究不同调控（不同植物生长调节剂、不同夏剪强度）处理的葡萄二次果花芽分化的生理和分子机制及成花情况，探索最优促进花芽分化的夏剪和植物生长调节剂处理方式，确保二次果成花稳定。研究结果可为稳定葡萄二次果产量提供实用的技术及理论依据。

随着一年两收栽培技术的推广应用，广西已经发展成为冬葡萄栽培特殊优势产区。但是部分品种二季果成花较差，一定程度上阻碍了葡萄一年两收栽培技术的进一步推广。为明确葡萄二季果成花的生理分子机制和调控机理，本研究以8年生一年两收栽培的夏黑葡萄为试材，进行不同摘心强度和喷施不同生长调节剂处理，通过制作石蜡切片观察统计花芽形态分化进程及成花率。同时测定了不同时期冬芽中植物内源激素的含量和花分生组织特征基因的表达情况。并利用生物信息学和RNA-seq技术，全面解析了矮壮素（CCC）处理对二季果冬芽分化关键时期差异基因表达水平的影响。具体结果如下：.(1)摘心处理可以促进一年两收夏黑葡萄花芽分化，缩短花芽分化进程时间，提高成花率。花上2片叶、4片叶、6片叶摘心处理的各节位进行花芽分化的芽从进入花原始体发育期到最先完全进入或通过花穗形成期时经历的时间比对照相应节位缩短了15-31 d，成花率比对照提高了13.3%-46.7%。喷施不同浓度的6-BA可以缩短花芽分化进程时间8-23 d，提高成花率6.7%-20.0%，其中以对第6、7节位的芽影响最好；喷施CCC处理也可以缩短花芽分化进程和促进成花，但效果不如6-BA。而喷施ETH可以缩短花芽分化进程时间但对成花率无明显促进作用。.（2）夏黑葡萄中8个花分生组织特征基因的表达受到摘心和CCC处理的影响，其中从-3 DAF到11 DAF，较高水平的FT、TFL1A和TFL1B的表达及较低水平的LFY基因的表达会促进夏黑葡萄二季果花序原基的形成，18 DAF时较高水平的CAL A、SOC1和TFL1A的表达会促进花序原基的发育。-3和4 DAF时，摘心和CCC处理植株中ZR/GAs比值较低，会促进始原基的形成。.（3）构建了CCC处理后夏黑葡萄二季果冬芽分化关键时期的RNA-seq文库，发现在-3和4 DAF，CCC处理会降低大多数差异基因的表达量，而在11 DAF，CCC处理后上调表达的差异基因数目较多。CCC处理在4 DAF时对二季果冬芽内基因表达水平的影响最为显著，筛选并鉴定了响应CCC处理的与苯丙氨酸代谢、苯丙素生物合成、类黄酮生物合成、激素、转录因子及成花相关的差异表达基因。