高温胁迫下萝卜苗期的转录组分析

Transcriptome analysis of Raphanus sativus seedlings in response to high temperature stress

  • 摘要: 【目的】对高温胁迫下萝卜幼苗进行转录组测序分析,筛选不同耐热性萝卜品种间的差异表达基因,为阐明萝卜幼苗响应高温胁迫的分子机制提供理论参考。【方法】以耐热萝卜品种1116T和热敏感萝卜品种Wr129为对象,对其进行高温(40℃)胁迫处理0(常温,对照)和24 h,提取各时间点样品总RNA进行转录组测序,以|log2 FoldChange|≥1,P<0.05且TPM≥10为标准,筛选出差异表达基因(DEGs),并进行GO功能注释和KEGG代谢通路富集分析。【结果】随着高温胁迫时间的增加,Wr129和1116T幼苗茎叶逐渐发生萎蔫,胁迫24 h后Wr129叶片萎蔫下垂且干枯变黄,而1116T叶片轻度萎蔫下垂,仍保持绿色。12个文库共获得81.45 Gb Clean data,平均有62.30%的reads比对到萝卜参考基因组的一个位置。从2个不同耐热性品种中共鉴定到2701个差异表达基因。GO功能注释结果显示,2629个差异表达基因在GO数据库中得到注释,其中,有559个差异表达基因被注释为细胞组分类别中的叶绿体被膜、叶绿体基质和叶绿体类囊体3个条目,470个差异表达基因被注释为与胁迫响应相关的生物学过程。KEGG代谢通路富集结果显示,1166个差异表达基因富集到124条代谢通路,其中,有74个差异表达基因富集到光合作用和光合作用-天线蛋白代谢通路,23个基因富集到卟啉和叶绿素代谢通路,大部分基因在高温胁迫下被诱导; 35个差异表达基因富集到谷胱甘肽代谢途径,其中14个谷胱甘肽-S-转移酶基因(GSTs)在高温胁迫下上调表达。高温胁迫24 h,Wr129和1116T幼苗叶片PSII的最大光化学效率(Fv/Fm)和总叶绿素含量均下降,但1116T的下降幅度较小。【结论】 Wr129和1116T的耐热性存在明显差异,其原因是1116T植株可通过调控光合作用和抗氧化系统相关基因的表达,减缓Fv/Fm和叶绿素含量下降速率,从而提高植株对高温胁迫的耐受能力。

     

    Abstract: 【Objective】To investigate the difference of gene expression in Raphanus sativus seedlings responding to the high temperature through transcriptome sequencing analysis, so as to provide theoretical basis for the molecular mechanism of tolerance to heat stress in R. sativus.【Method】 The heat-resistant radish variety 1116T and heat-sensitive radish variety Wr129 were treated with high temperature(40℃)stress for 0(room temperature,control)and 24 h. The total RNA of samples at each time point was extracted for transcriptome sequencing. Taking|log2 Fold Change| ≥ 1,P<0.05 and TPM ≥ 10 as the standard,the differentially expressed genes(DEGs)were screened,and GO function and KEGG metabolic pathway enrichment analysis were performed.【Result】 With the increase of heat stress time, the stems and leaves of Wr129 and 1116T seedlings gradually wilted. After 24 h of stress, the leaves of Wr129 wilted,drooped and became dry and yellow, while the leaves of 1116T were slightly wilted, drooped and remained green. A total of 81.45 Gb clean data were obtained from 12 libraries, and on average of 62.30% of the reads were mapped to the reference genome. In total, 2701 DEGs were screened. Through GO functional annotation analysis, 2629 DEGs were annotated in the GO database, of which 559 DEGs were enriched to three items of chloroplast envelope, chloroplast stroma and chloroplast thylakoid in cellular component categories, and 470 DEGs were annotated as related to stress response in biological process. KEGG metabolic pathway enrichment showed that 1166 DEGs were annotated into 124 metabolic pathways, of which 74 DEGs were enriched in photosynthetic pathway and the photosynthesis-antenna protein pathway, 23 DEGs were enriched in the porphyrin and chlorophyll metabolic pathway, and most genes were induced under heat stress. While 35 DEGs involved in the glutathione metabolism pathway, and 14 glutathione-S-transferase genes(GSTs)up-regulated in this pathway. The maximum photochemical efficiency(Fv/Fm)and total chlorophyll content of PSII in leaves of Wr129 and 1116T seedlings decreased under high temperature stress for 24 h,but the decrease of 1116T was small.【Conclusion】 The heat tolerance of Wr129 and 1116T is obviously different, because 1116T can improve its heat resistance by the regulation of metabolic process relating to antioxidant and photosynthetic systems and slow the Fv/Fm value and chlorophyll content rate.

     

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