烟草响应铬胁迫microRNA与蛋白质的鉴定

发布时间：2018-04-23 02:22

本文选题：铬 + 基因型差异　；参考：《浙江大学》2015年博士论文

【摘要】：土壤重金属污染是对人类健康构成威胁的一个重要环境因素。烟草作为重要的经济作物,在叶片中能够积累一定量的重金属,危害人类健康。六大有毒的重金属中,铬(Cr)排第二位,主要通过人类活动被释放到环境中。六价态铬有毒且易诱变致癌,因此迫切需要阐明植物耐铬机制,培育高耐铬性但铬低积累的烟草品种。本研究以耐铬性不同烟草基因型为材料,旨在探讨铬胁迫对烟草生长与生理特性的影响及基因型差异,分离鉴定耐铬相关的miRNAs和蛋白质。主要研究结果如下： 1.铬胁迫对烟草生长、矿质元素含量、超微结构和光合特性的影响及基因型差异以两个耐铬性不同的烟草基因型贵烟1号(耐铬基因型)和云烟2号(铬敏感基因型)为材料,温室水培铬胁迫试验,研究铬胁迫对烟草植株生长、超微结构、矿质元素含量和光合特性的影响。结果表明,100μM铬胁迫显著抑制烟草植株的生长,降低株高、根长和生物量,其中铬敏感基因型云烟2号比耐铬基因型贵烟1号受抑制更严重。铬胁迫降低叶片叶绿素含量、净光合速率(Pn)和光系统Ⅱ的最大光化学效率(Fv/Fm)；同时,显著减少根和叶片中的Zn含量以及根中的Fe和Mn含量,但提高叶片中的Fe含量。透射电子显微镜观察结果表明贵烟1号比云烟2号受铬胁迫的影响较小,贵烟1号叶片表现相对较好的光合器官组成结构。 2.烟草根系中铬胁迫响应niRNA和目标基因的全基因组鉴定为了在整体水平上系统地分析铬响应miRNA及其目标基因,以两个耐铬性不同的烟草基因型贵烟1号和云烟2号为材料,温室水培铬胁迫试验,以无铬基本培养液为对照,建立根系四个niRNA文库：贵烟1号-铬处理、贵烟1号-对照、云烟2号-铬处理和云烟2号-对照。利用高通量测序技术,鉴定得到53个保守的和29个新的miRNA家族。对41个保守的铬响应miRNA家族进行比较基因组分析,结果表明11个miRNA家族在贵烟1号中上调但在云烟2号中不变,17个miRNA家族只在铬胁迫下的云烟2号中上调。只有一个nmiRNA家族miR6149,在云烟2号中下调但在贵烟1号中不变。29个新的niRNA家族中,铬胁迫下有14个在两个基因型中差异表达。通过高通量降解组测序同源性搜索,对41个保守的和14个新的铬响应miRNA家族的目标基因进行预测。COG功能类别分析结果表明一些预测的miRNA目标转录体对生物和非生物胁迫有响应。另外,通过RT-PCR验证了部分铬响应miRNA的表达模式。试验结果为理解miRNA在铬耐性中的功能提供了有价值的信息和框架。 3.铬胁迫对烟草叶片蛋白表达谱的影响及基因型差异利用比较蛋白质组学技术,探讨了铬胁迫对贵烟1号和云烟2号叶片蛋白质表达谱的影响及基因型差异,与对照相比,100μM铬胁迫下贵烟1号共有86/74个蛋白上调/下调,云烟2号共有107/77个蛋白上调/下调。鉴定到12个耐铬相关蛋白,100μM铬胁迫下这些蛋白在贵烟1号中显著表达但在云烟2号中下调或不变,或者在贵烟1号中不变但在云烟2号中下调,上述耐性相关蛋白包括线粒体肽酶、脱水蛋白、超氧化物歧化酶、腺嘌呤转磷酸核糖基酶和线粒体苹果酸脱氢酶等；蛋白功能分类分别属于胁迫响应、能量代谢、RNA结合和代谢相关蛋白。
[Abstract]:The heavy metal pollution in soil is an important environmental factor which threatens human health . As an important economic crop , it can accumulate a certain amount of heavy metals and harm human health . In six poisonous heavy metals , chromium ( Cr ) is released into the environment .

1 . Effects of chromium stress on tobacco growth , mineral element content , ultrastructure and photosynthetic characteristics and genotype difference

The effects of chromium stress on growth , ultrastructure , mineral element content and photosynthetic characteristics of tobacco plants were studied . The results showed that chromium stress inhibited the growth of tobacco plants , reduced plant height , root length and biomass .
At the same time , the content of Zn in roots and leaves and the content of Fe and Mn in roots were significantly reduced , but the content of Fe in the leaves was increased . The results of transmission electron microscopy showed that the effect of No . 1 was smaller than that of Yunyan No . 2 , and the leaves of Guido No . 1 showed relatively good composition structure of photosynthetic organs .

2 . Complete genome identification of the response of chromium stress response to niRNA and target gene in tobacco root system

In order to systematically analyze the chromium response miRNA and its target gene at the whole level , we identified four conservative and 29 new miRNA families by using high - throughput sequencing technology . The results showed that 11 miRNA families were up - regulated in Guiyan No . 1 , but there were 14 new miRNA families in Yunyan No . 1 .

3 . Effect of chromium stress on protein expression profile and genotype difference of tobacco leaf protein

The effects of chromium stress on the protein expression profile of No . 1 and Yunyan No . 2 leaves were investigated by comparative proteomic techniques . Compared with the control group , there were 86 / 74 proteins up - regulated / down - regulated under 100 渭M chromium stress , and there were 107 / 77 proteins up - regulated / down - regulated in Yunyan No . 2 .
Protein functional classification belongs to stress response , energy metabolism , RNA binding and metabolism - related proteins , respectively .

【学位授予单位】：浙江大学
【学位级别】：博士
【学位授予年份】：2015
【分类号】：S572

【参考文献】