玉米光周期敏感近等基因系的转录组分析

发布时间：2018-05-06 19:58

本文选题：玉米 + 光周期敏感性　；参考：《河南农业大学》2016年硕士论文

【摘要】：玉米(Zea mays ssp.mays L.)是世界上分布最广泛的粮食作物之一,近年来在我国的种植面积超越小麦和水稻而居第一位。为了发现更多新的玉米品种,热带亚热带玉米凭借着丰富的遗传变异,成为最主要的研究对象。但在温带地区这些品种的光周期的敏感性成为制约品种选育的主要障碍。至今,玉米光周期的研究已经被初步报道,但是光周期路径中的相关基因具体是通过哪些机制来影响玉米的开花发育,尚不清楚。植物光周期的过程极其复杂,涉及众多基因的协调参与,本研究以温带光周期不敏感的黄早4(H4)以及与光周期敏感热带自交系CML288构建的近等基因系496-10(H4-NIL)为试验材料,在长日照条件下不同时期的茎尖观察分析光周期敏感时期,并以不同时期的叶片进行转录组高通量测序,结合生物信息学、RT-PCR等方法,分析两种材料在长日照条件下不同发育时期(营养生长期、营养生长向生殖生长转化期、生殖生长期)的表达模式及调控路径。本研究的主要结论如下:1.通过茎尖形态观察,发现H4的茎尖在3、4、5、6叶期分别与H4-NIL的茎尖在3、5、6、7叶期时有相一致的发育形态。根据茎尖形态将其定义为三个发育时期,营养生长期;营养生长向生殖生长转化时期;生殖生长期。利用RNA-seq技术对H4的3、4、5、6叶期及H4-NIL的3、5、6、7叶期进行高通量测序。得到的reads数在2500~2800万之间,这其中大约有64%的reads在基因组上有唯一的匹配位点。2.生物信息学分析表明,H4和H4-NIL的对应叶期分别有357、531、736、655个差异表达的基因,在上调的基因中,分别有99、126、94、203个基因在4个不同叶期特异表达,其中有4个基因在4个叶期中都上调;下调的基因中,分别有122、279、417、229个基因在4个不同叶期特异表达,其中12个基因在4个叶期都下调。根据GO注释,富集到的基因功能主要集中于催化活性、氧化还原酶活性、代谢过程和氧化还原反应四大类。而在生物过程类别中,这些差异基因主要参与离子传输和光合作用,利用Cytoscape(v3.0.2)插件ClueGO+Cluepedia v2.1.3,结合差异基因,生物过程以及KEGG通路分析,将该差异基因进行分类,我们发现很多基因涉及不仅一个功能类别,说明光周期敏感机制涉及多种功能及代谢调控,并可能与其他机制调控相互作用。3.通过差异相关基因聚类,按照不同叶期的表达量变化将这些差异表达基因分为三类:第一类可能是在营养生长阶段向生殖生长阶段转化的阶段起主要作用;第二类基因可能更多的参与了营养生长的调控;第三类可能与生殖生长阶段的生长发育调控有关。4.选取H4和H4-NIL的4个叶期存在表达差异的19个光周期相关基因,根据它们不同叶期的表达量变化将其分为三类:第一类基因GRMZM2G002220、GRMZM2G013398、GRMZM2G076602、GRMZM2G138750、GRMZM2G016756,在营养生长和生殖生长阶段上调,在营养生长向转化阶段下调;第二类AC233888.1_FG002、GRMZM2G062541、GRMZM2G156692、GRMZM2G449950、GRMZM2G021777、GRMZM2G092363、GRMZM2G162737、GRMZM2G165488,在营养生长向生殖生长转化阶段上调,在营养生长和生殖生长阶段下调;第三类GRMZM2G175265、GRMZM2G474769、GRMZM2G004483、GRMZM2G014902、GRMZM2G138455、GRMZM2G165042,在营养生长阶段上调,在营养生长向生殖生长阶段和生殖生长阶段下调。5.通过光周期差异基因间表达模式分析,说明CCA1/LHY、TOE3以及COL9的表达量较高引起CO表达量降低导致H4-NIL开花延迟。此外,这些光周期敏感差异表达基因还涉及种子萌发、幼苗生长以及对非生物胁迫的响应等。
[Abstract]:Zea mays ssp.mays L. is one of the most widely distributed grain crops in the world. In recent years, the planting area in China is the first to exceed the wheat and rice. In order to find more new maize varieties, the tropical and subtropical corn has become the most important research object with its rich genetic variation. The photoperiod sensitivity of the photoperiod has become a major obstacle to the selection of varieties. Up to now, the study of the Photoperiod of maize has been preliminarily reported, but it is not clear that the related genes in the photoperiod path affect the flowering and development of maize. The process of the plant photoperiod is extremely complex, involving the coordinated participation of many genes. In this study, the photoperiod insensitive yellow morning 4 (H4) and the near isogenic line 496-10 (H4-NIL), which were constructed with the photoperiod sensitive inbred line CML288, were used as experimental materials to observe the photoperiod sensitive period of the stem apex at different periods under long sunshine conditions, and the high throughput sequencing of the transcriptional groups at different periods was carried out in combination with biisin. The expression patterns and regulation paths of two kinds of materials in different developmental periods (vegetative growth period, vegetative growth and reproductive growth period, reproductive growth period) under long sunshine conditions were analyzed by means of RT-PCR and other methods. The main conclusions of this study are as follows: 1. the tip of the stem at the 3,4,5,6 leaf stage and the tip of the stem of H4-NIL were found at the leaf point of the stem at the leaf point of the stem. 3,5,6,7 has a consistent developmental morphology during the leaf stage. According to the morphology of the stem tip, it is defined as three developmental periods, vegetative growth period, vegetative growth transformation period to reproductive growth, reproductive growth period, and high flux sequencing of 3,4,5,6 leaf phase and H4-NIL 3,5,6,7 leaf phase of H4 by using RNA-seq technology. The number of reads is between 2500~2800 million, About 64% of the reads had the only matching site in the genome.2. bioinformatics analysis showed that there were 357531736655 differentially expressed genes in the corresponding leaf period of H4 and H4-NIL. In the up regulated genes, there were 99126,94203 genes at 4 different leaf stages respectively, of which 4 genes were all in the 4 leaf stages. In the down regulated genes, 122279417229 genes were expressed specifically at 4 different leaf stages, of which 12 genes were downregulated in the 4 leaf stages. According to the GO annotation, the enriched gene function mainly concentrated on the catalytic activity, the oxidoreductase activity, the metabolic process and the redox reaction in four major categories. In the biological process category, these differences were different. Different genes are mainly involved in ion transmission and photosynthesis, using the Cytoscape (v3.0.2) plug-in ClueGO+Cluepedia v2.1.3, combining differential genes, biological processes and KEGG pathway analysis to classify the differential genes. We found that many genes involve not only one functional category, but also that the photoperiod sensitive mechanism involves a variety of functions and metabolic modulation. Control, and may control interaction with other mechanisms,.3. can be classified into three categories according to the variation of different leaf stages, and the first class may play a major role in the stage of transformation from the vegetative stage to the reproductive stage, and the second genes may be more involved in the nutrition growth. Regulation of the third types of photoperiod related genes related to the regulation of growth and development of the reproductive growth stage related to the 4 leaf stages of.4. selection of H4 and H4-NIL, which are divided into three categories according to the changes in the expression of their different leaf stages: the first class GRMZM2G002220, GRMZM2G013398, GRMZM2G076602, GRMZM2G138750, GRMZM2G016756 The second types of AC233888.1_FG002, GRMZM2G062541, GRMZM2G156692, GRMZM2G449950, GRMZM2G021777, GRMZM2G092363, GRMZM2G162737, GRMZM2G165488 are up regulated in the phase of the vegetative growth and the reproductive growth stage. Third kinds of GRMZM2G175265, GRMZM2G474769, GRMZM2G004483, GRMZM2G014902, GRMZM2G138455, GRMZM2G165042, up regulated during the vegetative growth stage. The analysis of the expression pattern of.5. through the photoperiod difference between the vegetative growth stage and the reproductive stage of the reproductive growth stage was analyzed. It is said that the higher expression of CCA1/LHY, TOE3 and COL9 causes CO expression. In addition, these photoperiod sensitive differentially expressed genes also involve seed germination, seedling growth and response to abiotic stress in H4-NIL.

【学位授予单位】：河南农业大学
【学位级别】：硕士
【学位授予年份】：2016
【分类号】：S513

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