半野生棉对复合盐碱的响应机制与克劳茨基棉盐胁迫转录组分析

发布时间：2018-04-16 02:06

本文选题：半野生棉 + 盐碱　；参考：《华中农业大学》2017年博士论文

【摘要】：棉花是重要的纤维和能源作物,也是盐碱地种植的先锋作物。然而高浓度的盐胁迫依然会对其产量和质量造成影响。近年来,土壤盐渍化越来越严重,极大的制约了棉花的产量和质量。半野生棉和野生棉是经过严苛的自然界环境筛选进化的材料,其中半野生棉(Semi-wild cotton)是陆地棉的野生类型,又称陆地棉野生种系(Gossypium hirsutum race),遗传多样性丰富,同时半野生棉与栽培品种之间具有遗传亲和性,是研究棉花抗盐机理并进行抗性改良的良好材料。本研究制定了室内稳定鉴定半野生棉萌发期和苗期复合盐碱抗性的方法,并初步探讨了ROS在半野生棉对复合盐碱响应中的机制。采用转录组分析方法分析了克劳茨基棉中SOS、ROS、ABA相关基因参与高浓度盐胁迫的响应机制。主要研究结果如下:1、确立了室内鉴定半野生棉复合盐碱抗感性的方法:通过对温室鉴定结果与课题多年在新疆盐碱地块半野生棉的鉴定结果的一致性分析,确立了合适土样;通过对土样中主要离子成分的测定,明确了新疆次生盐碱地块中的主要成分,并实现了温室重现,在进行多次验证后,确认了方法的可行性;成功的筛选到稳定抗性材料玛利加朗特棉85和稳定敏感材料阔叶棉40,抗性对照材料中16和敏感对照材料中12。2、苗期半野生棉材料复合盐碱胁迫响应:表型与生理生化显示以上四种材料的抗性具有生理学差异,同时发现抗氧化相关的酶在抗感材料之间的活性差异。通过Illumina HiSeq?2500平台测序,对复合盐碱胁迫0h、3h、12h和48h的半野生棉玛利加朗特棉85、阔叶棉40与陆地棉对照材料中12和中16的叶片与根的转录组测序及生物信息学分析,发现64,737个基因在全部的样品中表达。差异基因分析发现在遭受复合盐碱胁迫后下调基因数目大于上调基因数目,GO富集分析发现,差异基因在响应氧化还原进程的条目中显著富集。3、活性氧相关酶类与基因在半野生棉复合盐碱胁迫响应中的机制:通过基因功能注释,发现共有265个ROS产生与清除相关基因。对4个材料的各自的共同差异基因与活性氧代谢相关的基因进行检索发现有中12(152),中16(124),阔叶棉40(144),玛利加朗特棉85(131)个活性代谢通路相关的基因在各个材料中参与ROS调控。通过SOD、POD、CAT和RBOH四个基因家族的表达谱分析,我们确定了主要参与活性氧通路的基因。4、野生棉克劳茨基棉盐胁迫响应机制与转录组分析:根据多个盐浓度处理后克劳茨基棉与戴维逊氏棉的表型比对,确定了克劳茨基棉的抗盐性。在对300 mM NaCl处理后的克劳茨基棉叶片进行生理生化指标测定时,发现ABA、GSSH、H2O2等含量迅速升高。进而通过Illumina HiSeq?2500测序平台,对300 mM NaCl胁迫0h,3h,12h,48h的叶片与根的转录组测序及生物信息学分析,发现37,278条基因和797条新基因表达。差异基因分析发现根部差异基因在3个时间节点都显著富集。通过基因表达模式分析,发现叶片中的差异基因大部分下调表达。GO注释发现差异基因主要富集在响应胁迫与离子转运的分类上,KEGG注释发现差异主要富集在激素信号转导与合成通路。5、克劳茨基棉高盐胁迫响应通路:通过对离子转运、活性氧、激素相关基因的表达谱分析发现,SOS、ROS、ABA在棉花相应盐胁迫时起到非常重要的作用,结合前人研究,初步绘制棉花在高盐胁迫早期的响应通路。
[Abstract]:Cotton is an important fiber and energy crops, saline land planting pioneer crops. However, high concentration of salt stress will affect its yield and quality. In recent years, more and more serious soil salinization, which greatly restricts the yield and quality of cotton. Cotton and semi wild wild cotton is through the harsh nature the evolution of environmental screening materials, including semi wild cotton (Semi-wild cotton) is a type of wild cotton, also known as wild species of Upland Cotton (Gossypium hirsutum race), genetic diversity, and between semi wild cotton and cultivars with genetic affinity, is to study the mechanism of salt tolerance of cotton and improve the resistance of the good material. This study developed a method of indoor identification of stable semi wild cotton at Germination and seedling stage of mixed salt alkali resistance, and to explore the ROS in semi wild cotton on composite saline in the response by rotating mechanism. Record group analysis Klotz Ki cotton SOS, ROS, ABA genes involved in the response mechanism of high concentration of salt stress. The main results are as follows: 1. The established method of indoor identification of semi wild cotton composite salt anti perceptual: Based on the results of greenhouse identification and consistency in the identification of the subject for many years in Xinjiang semi wild saline land cotton analysis, established a suitable soil sample; the determination of main ions in soil samples, the main components of Xinjiang secondary salinization land, and realize the greenhouse in return, after repeated verification, confirm the feasibility of the method; successful screening to stable resistance materials and stable cotton Mary gallante 85 the sensitive material of broad-leaved 40 cotton, 12.2 resistant materials and 16 control sensitive control material, semi wild cotton seedling composite salt stress response: phenotypic and physiological and biochemical display more than four kinds of material resistance With physiological differences, also found that the antioxidant enzyme activity in the resistant and susceptible differences between the materials. Through the Illumina HiSeq 2500 platform? Sequencing of the composite salt stress 0h, 3h, 12h and 48h semi wild cotton Mary gallante 85 cotton, cotton 40 broad-leaved and upland cotton were analyzed in 16 and 12 and leaves root transcriptome sequencing and bioinformatics materials, found that the expression of 64737 genes in all the samples. Analysis of differentially expressed genes found in the suffering of mixed salt alkali stress is greater than the number of up-regulated genes down regulated genes after GO enrichment analysis showed that the number of genes in response to redox process entries were enriched in.3, enzymes related to active oxygen with the gene in semi wild cotton composite salt stress response mechanism in the annotation of gene function, found a total of 265 genes related to ROS production and scavenging. Based on 4 common different materials with live The gene of oxygen metabolism related searches have been found in 12 (152), 16 (124), 40 (144), broad-leaved cotton Mary gallante cotton 85 (131) active metabolic pathway related genes involved in the regulation of ROS in all materials. Through SOD, POD, CAT and RBOH expression analysis four gene family, we identified the.4 gene involved in ROS pathway, Wild Cotton Gossypium klotzschanium Anderss salt stress response mechanism analysis and transcriptome: according to phenotypic comparison of Gossypium klotzschanium Anderss and Davison's cotton multiple salt concentration treatment, determine the salt resistance of Gossypium klotzschanium Anderss. In G. klotzschianum leaves 300 mM after NaCl treatment were determined by physiological and biochemical indexes, ABA, GSSH, H2O2 content increased rapidly. And then through the Illumina HiSeq? 2500 to 300 mM NaCl sequencing platform, 3h, 12h, 0h stress, 48h analysis of the leaf and root transcriptome sequencing and bioinformatics, 37278 797 new genes and gene expression. The difference of gene analysis showed that root genes in 3 time nodes are significantly enriched. Through the analysis of gene expression patterns, differentially expressed genes were found in the leaves of most down regulated expression of.GO annotation found differentially expressed genes were mainly enriched in response to stress and the classification of transport, KEGG notes were found mainly enriched in hormone signal transduction and.5 synthesis pathway, G. klotzschianum high salt stress response pathway: Based on ion transport, reactive oxygen species, spectrum analysis showed that the expression of hormone related genes SOS, ROS, ABA play a very important role in the cotton corresponding to salt stress, combined with previous research, preliminary draw the response pathway of early cotton under high salt stress.

【学位授予单位】：华中农业大学
【学位级别】：博士
【学位授予年份】：2017
【分类号】：S562

【参考文献】