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可变剪切参与茶树对草甘膦胁迫的分子响应

Glyphosate-induced Alternative Splicing in Genes of Tea Plant

  • 摘要:
    目的 在茶园生产中,喷施草甘膦等除草剂时会发生药液漂移,从而严重影响茶树生长并危害茶叶品质和产量,但茶树在草甘膦胁迫下基因转录水平的变化及响应机制还知之甚少。
    方法 本研究以国家级茶树良种‘舒茶早’为试材,叶片喷施草甘膦(2.50 kg ai·hm−2)后在第1 d和第5 d进行转录组测序,并对差异基因的可变剪切(alternative splicing,AS)事件进行分析和验证。
    结果 内含子保留的AS类型占比最多(16.51%),其次是3'端AS(13.58%)和5'端AS(8.43%),而外显子跳跃发生频率最低(6.75%),其余类型占比54.73%;两个时间点分别有1588条和1768条差异可变剪切基因(DAGs),其中788条为共有DAGs,DAGs主要富集在类黄酮生物合成、丙酮酸代谢等与草甘膦胁迫相关通路;进一步证实了CsSDH1、CsSDH2、CsPR、CsADH、CsHD-ZipCsMYB等基因发生AS的真实性,并且qRT-PCR验证了CsSDH1、CsSDH2CsMYB基因及其可变剪切体的表达水平与其转录组表达水平高度一致。
    结论 本研究鉴定了大量的草甘膦胁迫条件下发生AS的差异基因,并证实了其AS事件的真实性,为进一步深入探究茶树对草甘膦胁迫响应的分子机制提供理论参考。

     

    Abstract:
    Objective Gene transcription and response mechanism of a tea plant exposed to glyphosate by herbicide spray were studied.
    Method Transcriptome sequencing was performed on the nationally renowned tea cultivar Shuchazao (Camellia sinensis) one day and 5 d after a spray of glyphosate at the rate of 2.50 kg ai·hm−2 on the field. Alternative splicing (AS) as detected in the differentially expressed genes was used to determine the molecular response of the plants.
    Result After the herbicide application, 16.51% AS was retained at the highest level in the introns, which was followed by 13.58% on the 3' end and 8.43% on the 5' end. The percentage of the unchanged in the exons was the least at 6.75%, while all the remainders at 54.73%. One day after the spraying, 1588 differential AS genes (DAGs), and 5 d, 1,768, were detected on the plants. Of them, 788 were found at both sampling times and all in the flavonoid biosynthesis or pyruvate metabolism pathway related to glyphosate stress. Significantly, the AS in CsSDH1, CsSDH2, CsPR, CsADH, CsHD-Zip, and CsMYB were positively observed with the qRT-PCR analysis showing highly agreeable results between the expressions and the transcriptomes of CsSDH1, CsSDH2, CsMYB, and the AS.
    Conclusion Numerous DAGs appeared in the tea plants exposed to glyphosate. With the confirmation, study on the molecular mechanism of the stress response as well as means to alleviate the ill-effects on the growth, quality deterioration, and yield decline on tea at plantations due to the herbicide application are in order.

     

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