【作者】 刘芳；

【导师】 吕秀兰；

【作者基本信息】 四川农业大学 ， 果树学， 2018， 博士

【摘要】 葡萄(Vitis Vinifera L.)为葡萄科葡萄属植物,和苹果、柑橘、香蕉一起被称为世界四大水果。据联合国粮农组织统计,中国在2016年成为世界葡萄的第一生产大国,鲜食葡萄年产量达1020万吨,占世界葡萄产量的46.4%。近年来,四川、重庆和贵州等西南部省份葡萄种植面积逐渐增大,但这些地区是典型的高温多湿和寡日照地区,打破休眠的需冷量积累不足而导致的萌芽不完全和不整齐、营养生长旺盛、产量低等问题成为这些地区葡萄产业发展的瓶颈。因此,本研究以四川高温多湿寡日照地区避雨栽培条件下‘香悦’和‘夏黑’葡萄为材料,通过研究不同破眠处理对葡萄生物学特性、总叶绿素含量动态变化等方面的影响,开发并完善了葡萄冬芽破眠技术;研究了单氰胺处理后‘夏黑’葡萄冬芽显微结构和转录组,探索了单氰胺调控葡萄冬芽萌发的机理。以上研究为四川高温多湿寡日照地区葡萄冬芽破眠提供了实践和理论指导。通过本试验研究得到如下结果:1、四川高温多湿和寡日照地区避雨栽培‘夏黑’和‘香悦’葡萄冬芽休眠经历类休眠、内休眠和生态休眠三个阶段,10月中旬前后处于类休眠阶段,10月中旬到12下旬处于内休眠阶段,1月初进入生态休眠阶段。2、以破眠剂浓度、涂芽时间和刻芽为因素正交设计进行破眠试验,结果表明:不同破眠处理均对‘香悦’葡萄的物候期产生影响,使得‘香悦’葡萄冬芽萌发时间提前6~28 d,果实成熟时间提前6~36d;使得‘夏黑’葡萄冬芽萌发时间提前4~29d,果实成熟时间提前6~36d。分别用萌芽期和果实成熟期提前的时间作为两个品种葡萄破眠效果优劣衡量指标,分析本试验设定的破眠剂浓度、涂芽时间和是否刻芽各因素的影响程度,极差分析结果表明:涂芽时间对于打破葡萄冬芽休眠最为重要。通过对不同破萌处理的‘夏黑’和‘香悦’葡萄物候期观察、冬芽萌发率、成枝率和果枝率统计和果实品质等进行综合分析,表明1月中旬前后,2.5%-3.0%单氰胺涂芽结合刻芽处理对打破四川高温多湿和寡日照地区避雨栽培的‘香悦’和‘夏黑’葡萄休眠效果佳。‘香悦’葡萄萌芽率显著高于对照,‘夏黑’葡萄萌芽率、成枝率和果枝率均显著高于对照。果实品质与对照无显著差异,均表现为品种固有品质。3、在前期研究基础上,本研究还观察了2.5%单氰胺处理‘夏黑’葡萄冬芽分生组织显微结构和超显微结构的变化。结果发现单氰胺处理葡萄冬芽15 d时,分生组织显微结构和超微结构均表现出明显的变化。葡萄冬芽分生组织显微结构生长点逐渐增大并伸长,顶端逐渐变宽,细胞数量增加,生长点逐渐向上凸起,变宽变平,叶原基分化增多,芽主轴分化伸长,节间变宽,随着生长点的生长,不断生长新的叶原基;超微结构研究发现2.5%单氰胺处理‘夏黑’葡萄冬芽15 d时,细胞核体积增大、细胞器特别是线粒体、高尔基体和液泡的数量增加,液泡体积由大变小,及胞间连丝增粗和变多。4、以当年生‘夏黑’葡萄枝条在落叶后剪成的单芽枝段作为试验材料,取未处理0d、清水处理15 d和2.5%单氰胺处理15 d的休眠芽进行转录组测序。在测序结果中共检测到29551条基因,占指定参考基因组的73.29%,其中检测到660条新基因,通过与各数据库进行序列比对以及氨基酸序列的预测,共有545条基因获得功能注释。清水处理15 d和2.5%单氰胺处理15 d相比,检测到1774条差异基因,其中1090条上调,684条下调。KEGG分析发现这些差异基因主要集中在光合作用、淀粉与蔗糖代谢、脂肪酸代谢、植物激素信号转导和苯丙素生物合成等作用通路;GO功能注释这些差异基因主要涉及细胞过程、新陈代谢过程、氮化合物代谢过程、应急反应以及生物调控等。更多还原

【Abstract】 The grape(Vitis Vinifera L.)is a perennial deciduous woody vine fruit tree belonging to the genus Vitis,which is known as the world’s four major fruits together with apples,citrus and bananas.According to FAO statistics,China’s table grape production in 2016 was 10.2 million tons,accounting for 46.4% of the world’s total grape production that year,making it the world’s largest grape producer.In recent years,the planting area of grapes has gradually increased in southwestern provinces such as Sichuan,Chongqing and Guizhou,but these areas are typical areas of high temperature,high humidity,and low solar radiation.The problems of incomplete and irregular germination,vigorous nutrition growth and low yieldcaused by insufficient accumulation of cold demand for breaking dormancy have become the bottleneck of grape industry development in these areas.Therefore,in this study,grape under the conditions of rain shelter cultivation in high temperature,humidity and low solar radiationareas in Sichuan Province was used as the material.By studying the effects of different dormancy breaking treatments on the biological characteristics and the dynamic changes of total chlorophyll content of grape,the technical system of grape winter bud breaking dormancy was developed and improved.The microstructures and transcriptomes of winter buds of ’Summer Black’ grape treated with cyanamide were also studied,and the regulatorymechanism of the germination of winter buds was explored.The above studies provide practical and theoretical guidance for grape winter buddormancy breaking in high temperature,humidity and low solar radiation areas in Sichuan.The following results are obtained through this test study:1.Winter bud dormancy of grape cultivated in high temperature,humidity and low solar radiation areas of Sichuan Province experienced three stages of similar dormancy,internal dormancy and ecological dormancy.Around mid-October,it is the similar dormancy stage,and from mid-October to late-December,it is the internal dormancy stage.In early January,it enteres the ecological dormancy stage.2.The experiment was carried out by orthogonal design,taking coating agent,coating time and carved bud as factors.The results showed that different treatments had effects on the phenological period of ’Xiang Yue’ grape,the germination time of winter bud of ’Xiang Yue’ grape was 6~28 days earlier,and the ripening time of the fruit was 6~36 days earlier.The germination time of winter bud of ’Summer Black’ grape was 4~29 days earlier,and the ripening time of the fruit was 6~36 days earlier.The time advanced in germination stage and the fruit maturation stage respectively were used as the indexes to evaluate the effect of dormancy breakingof the two grape varieties.The effects of the concentration of coating agent,the time of coating and whether or not to carve the buds in this experiment were analyzed.The results of the range analysis showed that: That is,the time of bud coating is the most important to break the dormancy of grape winter bud.The phenological observation of ’Summer Black’ and ’Xiang Yue’ grape,germination rate,branch formation and fruit branch rate of winter bud,and fruit quality of grape treated with different breaking treatments were analyzed.The results showed that the good effect was to break the dormancy of ’Xiang Yue’ and Summer Black’ grape in high temperature,humidity and low solar radiationareas in Sichuan Province around mid January,with 2.5%-3.0% cyanamide coating combined with bud carving treatment.The germination rate of ’Xiang Yue’ grape was significantly higher than that of the control.The germination rate,the branch formation rate and the fruit branch rate of Summer Black’ grape was significantly higher than that of the control.There were no significant difference between the fruit quality and the control of ’Xiang Yue’ and ’Summer Black’ grape,all of them are the inherent quality of the variety.3.On the basis of previous studies,the changes of microstructure and ultrastructure of winter bud meristem of ’Summer Black’ grape with 2.5% cyanamide treatment were also observed.The results showed that the microstructure and ultrastructure of the meristem showed obvious changes when treated with cyanamide for 15 days,but the change was not visible to the naked eye.The microstructural growing point of the meristem of winter bud of grape gradually increased and elongated,the top gradually widened,the number of cells increased,and the growing point gradually protruded upward,widening and flattening.The differentiation of leaf primordium increased,the main axis of bud is differentiated and elongated,and the internodes became wider.With the growth of the growing point,new leaf primordium was constantly growing.The ultrastructural study also found that when treated with 2.5% cyanamide of ’Summer Black’ grape for 15 days,the volume of the cell nucleusincreased,the number of organelles,especially mitochondria,Golgi apparatus and vacuoles increased,the volume of vacuoles changed from large to small,and the plasmodesmatathickened and increased.4.Taking the singlebud branch cut from the branches of ’Summer Black’ grape after falling leaves of the year as the experimental materials,the dormant buds of treated for CK,treated with clear water for 15 days and treated with 2.5 % cyanamide for15 days were sequenced by transcriptome.A total of 29551 genes were detected in the sequencing results,accounting for 73.29% of the designated reference genomes.Among them,660 new genes were detected and 545 genes were functional annotated through sequence alignment and amino acid sequence prediction with each database.Compared treated with clear water for 15 days and treated with 2.5 % cyanamide for15 days,1774 DEGs were detected,1090 of which were up-regulated and 684 down-regulated.KEGG analysis showed these DEGs were mainly located in pathways such as photosynthesis,starch and sucrose metabolism,fatty acid metabolism,phytohormone signal transduction and phenylpropanoid biosynthesis.GO functional annotation showed these DEGs are mainly involved in cellular processes,metabolic processes,nitrogen compound metabolism,emergency response and biological regulation.更多还原