拟南芥微丝结合蛋白ADF1超表达构建及其在盐胁迫下的作用
发布时间：2018-08-31 19:53
【摘要】：土壤盐碱化是影响农作物产量的一个非常主要的因素。盐分过高可以使植物体内的离子失衡,造成渗透胁迫等一系列非生物胁迫,影响植物的正常生长。因此我们对植物耐盐机理的研究不仅能够丰富植物响应逆境过程的理论知识,其研究的结果还可以用于农业生产实践,解决培育高耐盐性的农作物的问题。本实验室前期研究发现,微丝结合蛋白基因AtADF1可以被盐胁迫诱导,AtADF1突变体造成了植物对盐敏感,阻碍了盐诱导的前期微丝解聚过程。研究结果表明,微丝结合蛋白基因AtADF1的缺失影响拟南芥响应盐胁迫的过程。本实验首先通过克隆、构建、鉴定的方法获得了 3个拟南芥超表达ADF1(AtADF1-OEs)纯合株系,然后以拟南芥野生型(WT)和2个纯合株系的AtADF1-OEs为实验材料,观察拟南芥野生型(WT)和AtADF1-OEs纯合植株在盐胁迫下的萌发率,生长情况和微丝的动态变化,测定盐胁迫下拟南芥AtADF1-OEs纯合植株中盐胁迫响应基因SOS1、SOS2和SOS3的相对表达量,从而揭示超表达拟南芥微丝结合蛋白基因ADF1在盐胁迫下的作用机制。主要结果如下:(1)通过克隆AtADF1基因,构建AtADF1超表达载体,从而得到了AtADF1的超表达植株。(2)对AtADF1的超表达植株进行筛选和鉴定,总共获得了 3个不同表达量的纯合植株 AtADF1-OEs。(3)在正常条件下,拟南芥野生型(WT)和AtADF1-OEs植株的萌发率基本相同。在盐胁迫下(NaCl处理),AtADF1-OEs植株的萌发率要高于WT。(4)在正常条件下,拟南芥野生型(WT)和AtADF1-OEs纯合植株的生长情况基本一致。在盐胁迫下(NaCl处理),AtADF1-OEs纯合植株的生长趋势要强于WT植株,主要表现为侧根数、叶面积和根长均相对变大。(5)在正常条件下,拟南芥野生型(WT)和AtADF1-OE植株的微丝都是聚合的,没有发生解聚。在盐胁迫下(NaCl处理),与WT相比,AtADF1-OE植株增强了微丝的解聚。(6)在正常条件下,AtADF1-OEs纯合植株中SOS1、SOS2和SOS3基因表达量与WT植株相比基本相同。在盐胁迫下(NaCl处理),WT植株中SOS1、SOS2和SOS3基因表达量都是升高的;AtADF1-OEs纯合植株中SOS2和SOS3基因的表达量都高于WT植株。综上所述,AtADF1基因超表达后,有利于盐胁迫下植物的萌发和生长过程,增强了盐诱导的微丝解聚过程,提高了盐诱导的SOS2和SOS3基因的表达,说明AtADF1基因超表达提高了拟南芥耐盐的能力。
[Abstract]:Soil salinization is a very important factor affecting crop yield. Excessive salinity can cause ion imbalance in plants, resulting in a series of abiotic stresses, such as osmotic stress, which affect the normal growth of plants. Therefore, our research on the mechanism of plant salt tolerance can not only enrich the theoretical knowledge of plant response to stress, but also can be used in agricultural production to solve the problem of cultivating crops with high salt tolerance. Our previous study found that the AtADF1 gene of microfilament binding protein could be induced by salt stress to induce AtADF1 mutant to make plants sensitive to salt and hinder the process of microfilament depolymerization induced by salt. The results showed that the deletion of microfilament binding protein (AtADF1) gene affected the response of Arabidopsis thaliana to salt stress. In this experiment, three Arabidopsis superexpressed ADF1 (AtADF1-OEs) homozygous lines were obtained by cloning, construction and identification, and then the wild-type (WT) and AtADF1-OEs of two homozygous strains were used as experimental materials. The germination rate, growth and microfilament dynamics of wild-type Arabidopsis (WT) and AtADF1-OEs homozygous plants under salt stress were observed. The relative expression of SOS1,SOS2 and SOS3 genes in Arabidopsis AtADF1-OEs homozygous plants were measured. The mechanism of overexpression of microfilament binding protein (ADF1) gene in Arabidopsis thaliana under salt stress was revealed. The main results are as follows: (1) by cloning the AtADF1 gene and constructing the AtADF1 overexpression vector, the superexpression plants of AtADF1 were obtained. (2) the superexpression plants of AtADF1 were screened and identified. Three homozygous plants AtADF1-OEs. (3) with different expression levels were obtained. Under normal conditions, the germination rates of wild-type (WT) and AtADF1-OEs plants in Arabidopsis thaliana were almost the same. The germination rate of AtADF1-OEs plants under salt stress (NaCl treatment) was higher than that of WT. (4) under normal conditions, the growth of wild type (WT) and AtADF1-OEs homozygous plants of Arabidopsis thaliana was basically the same. Under salt stress (NaCl treatment), the growth trend of AtADF1-OEs homozygous plants was stronger than that of WT plants, mainly showing that the number of lateral roots, leaf area and root length were relatively larger. (5) under normal conditions, the microfilaments of Arabidopsis wild-type (WT) and AtADF1-OE plants were polymerized. There was no depolymerization. Under salt stress (NaCl treatment), AtADF1-OE enhanced the depolymerization of microfilaments compared with WT. (6) under normal conditions, the expression of SOS1,SOS2 and SOS3 genes in AtADF1-OEs homozygous plants was almost the same as that in WT plants. Under salt stress (NaCl treatment), the expression of SOS1,SOS2 and SOS3 genes in the homozygous plants of AtADF1-OEs was higher than that in WT plants. In conclusion, the overexpression of AtADF1 gene was beneficial to the germination and growth of plants under salt stress, enhanced the process of microfilament depolymerization induced by salt, and enhanced the expression of SOS2 and SOS3 genes induced by salt. The results showed that the overexpression of AtADF1 gene enhanced the salt tolerance of Arabidopsis thaliana.
【学位授予单位】：沈阳农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2;Q945.78
本文编号：2216001
[Abstract]:Soil salinization is a very important factor affecting crop yield. Excessive salinity can cause ion imbalance in plants, resulting in a series of abiotic stresses, such as osmotic stress, which affect the normal growth of plants. Therefore, our research on the mechanism of plant salt tolerance can not only enrich the theoretical knowledge of plant response to stress, but also can be used in agricultural production to solve the problem of cultivating crops with high salt tolerance. Our previous study found that the AtADF1 gene of microfilament binding protein could be induced by salt stress to induce AtADF1 mutant to make plants sensitive to salt and hinder the process of microfilament depolymerization induced by salt. The results showed that the deletion of microfilament binding protein (AtADF1) gene affected the response of Arabidopsis thaliana to salt stress. In this experiment, three Arabidopsis superexpressed ADF1 (AtADF1-OEs) homozygous lines were obtained by cloning, construction and identification, and then the wild-type (WT) and AtADF1-OEs of two homozygous strains were used as experimental materials. The germination rate, growth and microfilament dynamics of wild-type Arabidopsis (WT) and AtADF1-OEs homozygous plants under salt stress were observed. The relative expression of SOS1,SOS2 and SOS3 genes in Arabidopsis AtADF1-OEs homozygous plants were measured. The mechanism of overexpression of microfilament binding protein (ADF1) gene in Arabidopsis thaliana under salt stress was revealed. The main results are as follows: (1) by cloning the AtADF1 gene and constructing the AtADF1 overexpression vector, the superexpression plants of AtADF1 were obtained. (2) the superexpression plants of AtADF1 were screened and identified. Three homozygous plants AtADF1-OEs. (3) with different expression levels were obtained. Under normal conditions, the germination rates of wild-type (WT) and AtADF1-OEs plants in Arabidopsis thaliana were almost the same. The germination rate of AtADF1-OEs plants under salt stress (NaCl treatment) was higher than that of WT. (4) under normal conditions, the growth of wild type (WT) and AtADF1-OEs homozygous plants of Arabidopsis thaliana was basically the same. Under salt stress (NaCl treatment), the growth trend of AtADF1-OEs homozygous plants was stronger than that of WT plants, mainly showing that the number of lateral roots, leaf area and root length were relatively larger. (5) under normal conditions, the microfilaments of Arabidopsis wild-type (WT) and AtADF1-OE plants were polymerized. There was no depolymerization. Under salt stress (NaCl treatment), AtADF1-OE enhanced the depolymerization of microfilaments compared with WT. (6) under normal conditions, the expression of SOS1,SOS2 and SOS3 genes in AtADF1-OEs homozygous plants was almost the same as that in WT plants. Under salt stress (NaCl treatment), the expression of SOS1,SOS2 and SOS3 genes in the homozygous plants of AtADF1-OEs was higher than that in WT plants. In conclusion, the overexpression of AtADF1 gene was beneficial to the germination and growth of plants under salt stress, enhanced the process of microfilament depolymerization induced by salt, and enhanced the expression of SOS2 and SOS3 genes induced by salt. The results showed that the overexpression of AtADF1 gene enhanced the salt tolerance of Arabidopsis thaliana.
【学位授予单位】：沈阳农业大学
【学位级别】：硕士
【学位授予年份】：2017
【分类号】：Q943.2;Q945.78
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1 刘宇昌;拟南芥微丝结合蛋白ADF1超表达构建及其在盐胁迫下的作用[D];沈阳农业大学;2017年
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