珙桐苞片发育阶段的生理特征及ASR基因功能分析

发布时间：2018-03-31 17:08

  本文选题：珙桐　切入点：苞片发育　出处：《中南林业科技大学》2017年硕士论文

【摘要】：珙桐(Davidia involucrata Baill.)是珙桐科孓遗植物,在地球上生活了近6000万年,是极珍贵的树种,具有很高的研究价值,西方对珙桐在园林上的应用有很高的评价。珙桐最具代表性的即是其圆形的头状花序和成对的白色苞片。苞片是植物在生殖发育中出现的一类特殊的器官,广义上将其定义为与花或花序相关的叶片,珙桐是研究苞片发育的理想材料,珙桐苞片不仅表型十分独特,而且相较于苞片和叶片同时期生长,苞片的生长速度远远超过同时萌发的叶片。本研究主要从苞片发育过程中的生理生化分析及形态学观察:采集苞片不同发育阶段的样品,记录苞片颜色,大小苞片生长数据和苞片生长过程中的叶绿素含量等相关生理指标,将珙桐苞片的生长发育阶段进行分段分析,重点分析各发育阶段珙桐苞片内源激素含量变化。参考实际观测结果和激素变化情况,结合本实验组已有的珙桐苞片转录组数据库挑选与珙桐苞片生长发育有关的ASR基因,提取总RNA,克隆相关基因并用原核蛋白表达其特异性蛋白。使用适合于珙桐苞片发育定量分析的内参基因,根据表达模式对叶片与苞片,及苞片不同发育阶段的样品进qRT-PCR定量分析,利用qRT-PCR验证相关基因的表达规律。实验结果如下:1珙桐苞片形态发育规律珙桐苞片的生长发育阶段可以主要分为Ⅳ个时期:在苞片Ⅰ期,珙桐苞片在形成初期呈绿色,苞片与叶片同在混合芽中,芽鳞未完全脱落,与普通叶片形态差别很小,色差很小。苞片Ⅱ期,珙桐苞片颜色逐渐与叶片在色彩上出现差异,颜色变浅,苞片快速生长。苞片Ⅲ期,苞片绿色已经基本退去,略微带浅黄色,苞片生长速度稍微放缓。苞片Ⅳ期的苞片已经到达成熟期,纯白色,到达苞片的最大尺寸,生长停止。2.珙桐苞片发育过程中主要内源激素的变化规律使用高效液相色谱对珙桐苞片不同发育过程中的内源激素含量的测定显示:在苞片与叶片的比较中发现,珙桐苞片中的内源激素含量远远高于珙桐叶片中的含量。珙桐苞片发育的整个过程中主要内源激素含量的变化总体上是不断增加的;GA3与ABA的含量在珙桐苞片发育过程中是先上升,到最后阶段稍微下降;而IAA与ZT的含量在珙桐苞片发育过程中呈上升趋势,在最后阶段达到最大值。3.珙桐苞片ABA激素与DiASR表达量的变化规律从珙桐苞片转录组数据库中挑选出在珙桐苞片中与ABA有关且高量表达的DiASR基因。通过对珙桐苞片发育过程中内源激素含量的测定发现:ABA激素含量随着苞片生长发育而稳步增长的同时,当到达苞片Ⅲ期时达到最大值(ⅢⅡⅣⅠ),珙桐苞片发育Ⅲ期与其它几期的ABA激素含量有显著性差异。根据珙桐苞片各发育阶段的荧光定量结果表明:苞片的DiASR基因表达量是花序的200倍、是叶片的110倍,DiASR基因在苞片Ⅲ期的表达量最高,DiASR在珙桐苞片发育Ⅱ期表达量剧增,到Ⅲ期达到最高值(ⅢⅣⅡⅠ)。可以看出ABA含量和调控ABA表达的DiASR基因的变化情况基本一致,都在苞片Ⅲ期达到最大值,在苞片Ⅰ期达到最小值。依据两者的数据分析。发现两者的表达水平具有显著相关性。
[Abstract]:Dovetree (Davidia involucrata Baill.) is Nyssaceae relic plants, life on earth for nearly 60 million years, is the most valuable species, has high research value and application of Western Davidia involucrata in landscape are very high appraisal. Its most representative is white bracts round its head inflorescence and paired. Bracts is in a special class of plant organs appeared in the reproductive development, in broad sense, which is defined as with flower or inflorescence related leaf, bract development of Davidia involucrata is ideal material, not only a unique phenotype of bract of Davidia involucrata, and compared with bracts and leaves at the same period of growth. The growth rate is far more than the leaf bracts also germination. This study mainly developed in the process of physiological and biochemical analysis and morphological observation from bracts: samples of different developmental stages of acquisition bracts, record bract color, size of bract growth data and bracts The related physiological indexes during the growth of chlorophyll content, the growth stage of Davidia bract segment analysis, focuses on the analysis of changes of endogenous hormones in different developmental stages of Davidia bracts. The actual observation results and hormonal changes, combined with the experimental group have Davidia transcriptome database selection and bract of Davidia involucrata bract growth ASR gene related to the extraction of total RNA, gene cloning and expression of the specific protein with prokaryotic protein. Suitable for use in Davidia involucrate bract developed reference genes for quantitative analysis, according to the expression pattern of leaf and bract, bract and quantitative analysis of qRT-PCR in different developmental stages of sample, expression of genes related to verification by qRT-PCR. The experimental results are as follows: 1 Davidia bract morphology development regularity of Davidia bract growth stage can be divided into 4 stages: in stage I of bract, bract of Davidia involucrata In the early formation of a green bracts and leaves, with mixed buds, bud scales is not completely detached, very small and ordinary leaf shape difference, color difference is very small. Bracts II, bract and leaf color gradually Dovetree in color difference, color shallow, bracts bracts of rapid growth. Bracts green, has receded slightly pale yellow bracts slightly slower. The growth rate of bract bract stage has reached maturity, the pure white, reaching the maximum size of bracts, determination of endogenous hormone content of growth cessation changes of endogenous.2. in the process of development of bract of Davidia involucrata using high performance liquid chromatography of Davidia bracts in the process of development of different display: in the bracts and leaves, the content of endogenous hormones in the bract of Davidia involucrata is far higher than the content in the leaves. The main content of Dovetree endogenous hormones in the whole process of development in the bract of Davidia involucrata The overall change is increasing; the content of GA3 and ABA in the development process of Davidia bracts is increased to the final stage decreased slightly; while the content of IAA and ZT increased in Davidia bracts during development, reached the maximum value of.3. ABA DiASR hormone and bract of Davidia involucrata expression quantity pick out in Davidia bracts in ABA and the high expression of DiASR gene from bract of Davidia involucrata transcriptome in the last stage. Through the determination of the contents of endogenous hormones in the process of development of Davidia involucrate bract found: ABA with growth hormone content of bracts and steady growth at the same time, when the maximum arrival bracts of third stage (II III IV I), there was significant difference in development stage and bract of Davidia involucrata ABA hormone content in several other period. According to the quantitative fluorescence in different developmental stages of bract of Davidia involucrata results: DiASR gene expression is the bract inflorescence 200 times, 110 times the amount of leaves, DiASR gene expression was highest in bracts III, DiASR in Davidia bract developed phase II expression increase, to reach the highest value (stage III IV II I). See the changes in the content of ABA and the regulation of ABA expression of DiASR gene can be basically the same, are bracts in stage III reached the maximum in the bract stage and reached the minimum. Based on the analysis of the data found. Their expression level has a significant correlation.

【学位授予单位】：中南林业科技大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：S792.99
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本文编号：1691657