野生大豆种质资源耐盐性评价及离子转运相关基因对其耐盐能力的影响
本文选题:野生大豆 切入点:多样性分析 出处:《山东师范大学》2017年硕士论文
【摘要】:人们在耕作过程中的不合理灌溉、施肥、自然灾害等因素使得全球的土地盐碱化日趋严重。大豆是我国主要的油料作物,但随着育种进程中的不断选择,栽培大豆遗传多样性日趋狭窄。野生大豆是栽培大豆的野生近缘种,并且含有很多优异基因,例如:耐盐、抗旱、耐寒、抗虫、多荚等,尤其在耐盐方面比栽培大豆有着显著的优势。本研究以327份野生大豆种质资源为材料,通过表型性状及SSR分析,筛选并建立野生大豆多样性群体,对该群体进行表型性状及抗逆性多样性分析。并对其中耐盐及盐敏感极端材料进行耐盐生理生化分析、研究耐盐相关基因的表达情况,解析野生大豆耐盐机理。利用大豆发状根体系,通过耐盐相关基因的过表达及CRISPR-cas9技术突变目的基因,研究耐盐相关基因功能,为大豆耐盐育种提供重要信息。主要研究结果如下:1.对327份野生大豆资源的主茎类型、叶形、叶长、叶宽、叶柄长度、荚长、荚宽、花色等农艺性状进行调查,发现这些性状存在丰富的多样性,但有些材料相似性很高。利用课题组以前筛选出的具有较高多态性的10对SSR引物对327份野生大豆进行SSR分析,去除表型及SSR标记一致的材料,最终建立了101份野生大豆的多样性群体。2.对101份野生大豆多样性群体进行遗传多样分析发现,主茎类型、叶长、叶宽、叶柄长度、叶面积、荚长、荚宽、单株籽粒重、苗期株高、干重、耐盐性等方面均表现出了丰富的多样性,变异系数0.18~0.79,多样性指数为1.73~2.05。3.对耐盐材料ST-335和盐敏感材料SS-736进行盐胁迫下生理生化分析,发现盐胁迫条件下,与盐敏感品种SS-736相比,耐盐品种ST-335的叶片相对含水率高,可溶性糖含量高;SOD、POD和CAT活性高;Na+/K+低;且KOR,NHX和NSCC基因表达量低,而SKOR和SOS1基因表达量高。可能是ST-335具有较高的抗离子毒害、渗透胁迫及抗氧化胁迫的能力,且离子转运相关的一些基因通过协同作用控制Na+及K+的吸收和外排,最终决定ST-335具有相对较高的耐盐能力。4.在大豆发状根中过表达SKOR和SOS1基因,提高了大豆复合体的耐盐能力;而过表达NSCC基因,提高了大豆复合体对高盐胁迫的敏感性。在野生大豆发状根中利用CRISPR/Cas9体系成功诱导NSCC基因发生突变,NSCC发生突变的发状根中的Na+/K+比例显著低于野生型。进一步说明这些离子通道相关基因的变化对野生大豆耐盐能力产生影响。该方法也为野生大豆耐盐相关基因功能研究提供重要的理论基础。
[Abstract]:People in the farming process unreasonable irrigation, fertilization, natural disasters and other factors make the global land salinization has become increasingly serious. Soybean is the main oil crop in China, but with the continuous selection in the process of soybean genetic diversity is becoming more and more narrow. Wild soybean is the wild relatives of cultivated soybean, and contains a lot of excellent genes, such as salt tolerance, drought resistance, cold resistance, insect resistance, multi pod, especially in salt tolerance than cultivated soybean has the remarkable superiority. In this study, 327 wild soybean germplasm, through the analysis of phenotypic traits and SSR screening, and the establishment of wild soybean population diversity, analyze the Phenotype Traits and resistance diversity of the group. And the salt tolerant and salt sensitive materials for extreme salt tolerance physiological and biochemical analysis, the expression of genes related to salt tolerance, salt tolerance mechanism of wild soybean using. Soybean hairy root system by mutation gene expression and CRISPR-cas9 gene related to salt tolerance, salt tolerance related gene function research, provide important information for salt tolerance in soybean breeding. The main results are as follows: 1. of the 327 wild soybean resources stem type, leaf shape, leaf length, leaf width, petiole length. Pod length, pod width, color and other agronomic traits were investigated and found that the rich diversity of these traits, but some material similarity is very high. The research group previously screened with high polymorphisms of 10 SSR primers of 327 wild soybean by SSR analysis, SSR marker phenotype and removal consistent material. The eventual establishment of 101 wild soybean population diversity.2. of 101 wild soybean population diversity of genetic diversity analysis showed that the main stem type, leaf length, leaf width, petiole length, leaf area, pod length, pod width, grain weight per plant, seedling High, dry weight, salt tolerance has showed the rich diversity, the coefficient of variation was 0.18 ~ 0.79, the diversity index was 1.73 ~ 2.05.3. of salt tolerant and salt sensitive material ST-335 material SS-736 under Salt Stress on physiological and biochemical analysis, found that under salt stress, compared with salt sensitive strains of SS-736. Leaves of salt tolerant cultivar ST-335 relatively high moisture content, high content of soluble sugar; SOD, POD and CAT activity; Na+/K+ and KOR, is low; the expression of NHX and NSCC gene is low, and the expression of SKOR and SOS1 gene may be high. ST-335 has high anti permeability ability ion toxicity, oxidative stress and stress and, some genes related ion transport through synergistic absorption and control of Na+ and the role of K+, ST-335 finally decided to have a relatively high salt tolerance of.4. over expression of SKOR and SOS1 genes in soybean hairy root, improve soybean complex salt tolerance; and The expression of NSCC gene increased soybean complex sensitivity to high salt stress induced by CRISPR/Cas9 system. The NSCC gene mutation in wild soybean hairy roots, NSCC mutation of hairy root in the proportion of Na+/K+ was significantly lower than that of wild type. Further changes of the ion channel genes related to salt tolerance of wild soybean effect. The method also provides important theoretical basis for research on wild soybean salt tolerance related gene function.
【学位授予单位】:山东师范大学
【学位级别】:硕士
【学位授予年份】:2017
【分类号】:S565.1
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