苦荞耐盐基因FtGST2的克隆及功能初步分析
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1.山西农业大学农学院;2.中国农业科学院作物科学研究所

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国家自然科学基金(32161143005)


Cloning and Preliminary Functional Analysis of Salt Tolerance Gene FtGST2 in Tartary Buckwheat
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1.College of Agriculture,Shanxi Agricultural University;2.Key Laboratory of Grain Crop Genetic Resources Evaluation and UtilizationMARA,Institute of Crop Sciences,Chinese Academy of Agricultural Sciences;3.Chinese Academy of Agricultural Sciences

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National Natural Science Foundation of China(32161143005)

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    摘要:

    前期通过全基因组关联分析我们获得了一个苦荞耐盐相关基因FtGST2(FtPinG0707941400.01)。为了解FtGST2的基因结构与类型,我们通过同源比对和保守序列分析发现,苦荞中有69个GST基因家族的成员,该基因属于谷胱甘肽-s-转移酶(Glutathione S-transferases)中的四氯代氢醌脱卤素酶亚族(TCHQD)。对该基因上游2000 bp的启动子序列进行分析发现,34个元件中含有2个脱落酸(ABA)响应元件与6个茉莉酸甲酯(Me-JA)响应元件,而脱落酸与茉莉酸正是植物调控盐胁迫途径的重要激素。为进一步验证FtGST2的功能,我们从中苦3号中克隆出了FtGST2基因,对FtGST2在苦荞不同器官中的表达量和不同浓度NaCl处理下的表达量差异进行实验分析。结果显示,FtGST2在根中的表达量最高,并且FtGST2在100 mM NaCl处理下的表达量明显高于50 mM NaCl与未处理下的表达量。同时我们构建了FtGST2的过表达拟南芥与毛状根,并检测转基因材料在盐处理下的种子发芽率、主根长、毛状根鲜重及其生理指标。结果显示,过表达FtGST2的拟南芥发芽率和根长都高于与其相同处理的野生型,FtGST2的过表达毛状根在未处理条件下与对照组(A4)无明显差异,而在100 mM NaCl处理下的毛状根鲜重显著高于对照A4。对过表达毛状根与A4毛状根在100 mM NaCl处理下不同时间的表达量进行检测,发现FtGST2基因在受到盐胁迫3 h处表达量最高,6 h处下降,到12 h又有所上升。此外,对毛状根的生理指标进行检测后发现过表达FtGST2基因的确能有效提高苦荞毛状根过表达株系的耐盐性。以上实验对FtGST2基因的抗盐功能进行了初步验证,为后续苦荞耐盐品种的选育奠定了一定基础。

    Abstract:

    In the early stage, we obtained a salt-tolerant gene FtGST2(FtPinG0707941400.01) from tartary buckwheat by genome-wide association analysis, In order to understand the gene structure and type of FtGST2, we found that there were 69 members of the GST gene family in Tartary buckwheat, which belonged to the TCHQD in Glutathione S-transferases. Analysis of the 2000 bp promoter sequence upstream of the gene showed that 35 elements contained 2 ABA response elements and 6 Me-JA response elementsand abscisic acid and jasmonic acid were important hormones for plant regulation of salt stress pathway. In order to further verify the function of FtGST2, we cloned the FtGST2 gene from Zhongku-3, and the expression of FtGST2 in different organs of Tartary buckwheat and the differences in the expression of FtGST2 under different concentrations of NaCl treatment were analyzed. The results showed that the expression level of FtGST2 was the highest in roots, and the expression level of FtGST2 under 100 mM NaCl treatment was significantly higher than that under 50 mM NaCl treatment and no treatment. At the same time, we constructed Arabidopsis thaliana and hairy roots overexpressing FtGST2, and detected the seed germination rate, taproot length, hairy root fresh weight and physiological indexes of transgenic materials under salt treatment. The results showed that the germination rate and root length of Arabidopsis thaliana overexpressing FtGST2 were higher than those of the wild-type with the same treatment, and there was no significant difference between the hairy roots overexpressed by FtGST2 and the control group (A4) under the untreated conditions, while the fresh weight of hairy roots under 100 mM NaCl treatment was significantly higher than that of the control A4. The expression levels of overexpressed hairy roots and A4 hairy roots at different times under 100 mM NaCl treatment showed that the expression level of FtGST2 gene was the highest at 3 h after salt stress, decreased at 6 h, and increased at 12 h. In addition, the physiological indexes of hairy roots were detected, and it was found that the overexpression of FtGST2 gene could effectively improve the salt tolerance of tartary buckwheat hairy root overexpression lines. The above experiments preliminarily verified the salt resistance function of FtGST2 gene, which laid a certain foundation for the subsequent breeding of salt-tolerant varieties of tartary buckwheat.

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  • 收稿日期:2024-02-19
  • 最后修改日期:2024-04-12
  • 录用日期:2024-08-19
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