林泓, 黄子娟, 邓瑞莲, 等. NaCl胁迫下新型四倍体水稻种子萌发特性及基因差异表达分析[J]. 华南农业大学学报, 2023, 44(5): 696-707. DOI: 10.7671/j.issn.1001-411X.202306006
    引用本文: 林泓, 黄子娟, 邓瑞莲, 等. NaCl胁迫下新型四倍体水稻种子萌发特性及基因差异表达分析[J]. 华南农业大学学报, 2023, 44(5): 696-707. DOI: 10.7671/j.issn.1001-411X.202306006
    LIN Hong, HUANG Zijuan, DENG Ruilian, et al. Analysis of germination characteristics and differential gene expression in neo-tetraploid rice under NaCl stress[J]. Journal of South China Agricultural University, 2023, 44(5): 696-707. DOI: 10.7671/j.issn.1001-411X.202306006
    Citation: LIN Hong, HUANG Zijuan, DENG Ruilian, et al. Analysis of germination characteristics and differential gene expression in neo-tetraploid rice under NaCl stress[J]. Journal of South China Agricultural University, 2023, 44(5): 696-707. DOI: 10.7671/j.issn.1001-411X.202306006

    NaCl胁迫下新型四倍体水稻种子萌发特性及基因差异表达分析

    Analysis of germination characteristics and differential gene expression in neo-tetraploid rice under NaCl stress

    • 摘要:
      目的 评价新型四倍体水稻在不同浓度NaCl处理下的种子萌发特性、幼苗建成期形态指标和根部细胞结构差异,评估盐胁迫相关基因的表达差异。
      方法 在不同浓度NaCl (0、50、100、150、200和250 mmol/L)处理下,以‘华多1号’为研究材料,统计其种子在萌发时间、发芽势、发芽率和萌发指数指标的差异。在幼苗形成期,统计和比较‘华多1号’在不同浓度NaCl处理下的总根数、幼苗苗长、根长、幼苗鲜质量、含水率和根冠比参数的差异。利用WE-CLSM和塑料半薄切片技术观察根尖组织细胞的形态。利用基因组重测序、生物信息学分析和qRT-PCR技术对‘华多1号’与双亲相关的盐胁迫基因进行分析和定量验证。
      结果 当NaCl浓度升高时,‘华多1号’种子发芽率和萌发指数呈递减趋势,但相对盐害率和平均发芽时间呈递增趋势。当NaCl浓度高达250 mmol/L时,‘华多1号’种子相对盐害率为35%,表明其具有较强的耐盐性。在NaCl处理8~20 d后,‘华多1号’幼苗随着NaCl浓度升高,总根数呈减少趋势,同时幼苗苗长、根长、幼苗鲜质量和含水率呈下降趋势,但根冠比呈上升趋势。NaCl胁迫会抑制‘华多1号’幼苗生长,导致其根尖细胞体积变小,细胞皱缩和细胞排列松散;盐胁迫条件下,根尖伸长区的表皮和外皮层的细胞会增厚,中间皮层的细胞层数增加,中柱直径缩小,木质部导管分化数目增加。在‘华多1号’中发现有85个盐胁迫基因在CDS区中较双亲存在差异,包括53个盐敏感相关基因、32个盐胁迫抗性相关基因。qRT-PCR结果表明:与对照相比,‘华多1号’的4个盐敏感基因(LOC_Os03g16900LOC_Os06g48510LOC_Os02g34810LOC_Os04g32920)和2个耐盐性基因(LOC_Os03g20090LOC_Os11g26790)表达量显著上升。
      结论 该研究可为后续系统评价四倍体水稻种质资源的耐盐性、揭示四倍体水稻的耐盐性调控机理,以及后续筛选耐盐多倍体水稻品种提供理论依据。

       

      Abstract:
      Objective The purpose of this research was to evaluate differences in seed germination characteristics, morphological indexes, and root cell structure in ‘Huaduo1’ under different concentrations of NaCl and to evaluate the gene expression differences of salt stress related genes in ‘Huaduo1’.
      Method Using ‘Huaduo1’ as the research material, the differences in seed germination time, germination potential, germination rate, and germination index under various concentrations of NaCl (0, 50, 100, 150, 200, and 250 mmol/L) were analyzed. The differences in total root number, seedling length, root length, seedling fresh weight, water content, and root-to-head ratio of ‘Huaduo1’ at seedling forming stage under various concentrations of NaCl were summarized and compared. Root tip cell morphology of ‘Huaduo1’ was observed by the WE-CLSM and semi-thin section analysis. Genomic resequencing, bioinformatics analysis, and qRT-PCR analysis were conducted to analyze and quantify the expression of the salt stress genes in ‘Huaduo1’ related to its parents.
      Result The seed germination rate and germination index of ‘Huaduo1’ decreased with increased NaCl concentration, while the relative salt damage rate and average germination time increased. ‘Huaduo1’ showed a strong salt tolerance with the relative salt damage rate reached 35% under 250 mmol/L NaCl treatment. After 8 to 20 days of NaCl treatment, the total root number of ‘Huaduo1’ seedlings decreased with increased NaCl concentration. Meanwhile, the seedling length, root length, fresh weight and water content of seedlings under NaCl stress showed a decreasing trend, and the root-to-head ratio showed an increasing trend. NaCl stress inhibited the growth of the ‘Huaduo1’ seedlings, resulting in smaller cell volume, cell shrinkage, and loose cell arrangement in the root tip elongation zone. Moreover, the cells of the epidermis and outer cortex layer in the root tip elongation zone were thickened, the number of cell layers in the mesocortex increased, the diameter of the middle column decreased, and the number of xylem ductal differentiation increased under salt stress. Totally 85 salt stress-related genes of ‘Huaduo1’ showed variance in the CDS region compared with its two parents, including 53 salt sensitivity related genes and 32 salt stress tolerance related genes. The qRT-PCR results showed that the expressions of four salt sensitivity genes (LOC_Os03g16900, LOC_Os06g48510, LOC_Os02g34810, and LOC_Os04g32920) and two salt tolerance genes (LOC_Os03g20090 and LOC_Os11g26790) were significantly increased compared with the control.
      Conclusion The study can be used to evaluate the salt tolerance of tetraploid rice germplasm, reveal the regulation mechanism of salt tolerance in tetraploid rice, and provide a theoretical basis of the salt-tolerant germplasm selection in polyploidy rice.

       

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