沙芥幼苗叶片小热激蛋白基因的克隆与表达
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国家自然科学基金项目(31860553、31260475)


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    摘要:

    在对转录组和蛋白组数据联合分析的基础上,通过RT–PCR方法克隆得到沙芥幼苗叶片2个小分子热激蛋白基因PcHSP26.5和PcHSP17.8,运用生物信息学软件分析它们的核苷酸和编码蛋白,通过Real–time PCR分析其在高温、低温、NaCl、ABA和干旱复水胁迫下的表达模式。PcHSP26.5和PcHSP17.8开放阅读框分别为699 bp和462 bp,分别编码282和153个氨基酸,其中PcHSP26.5包含1个内含子。PcHSP26.5、PcHSP17.8的相对分子质量分别为26 480、17 490,理论等电点分别为6.36、5.99;均无跨膜结构与信号肽位点,属于亲水蛋白。亚细胞定位预测和进化树分析表明,PcHSP26.5主要定位于线粒体中,属于MⅡ亚族;PcHSP17.8主要定位于细胞质中,属于CⅠ亚族。实时荧光定量PCR结果表明:高温胁迫下,沙芥PcHSP26.5和PcHSP17.8基因的相对表达量均在1 h达到顶峰,之后下降;低温胁迫下,沙芥PcHSP26.5基因的相对表达量在0.5 h达到顶峰,之后下降,沙芥PcHSP17.8基因的相对表达量在0.5~12 h内低于对照,24 h时出现上升趋势;NaCl胁迫下,沙芥PcHSP26.5和PcHSP17.8基因的相对表达量分别在 1 h和3 h达到顶峰,之后下降;ABA胁迫下,沙芥PcHSP26.5和PcHSP17.8基因的相对表达量均在2 h达到顶峰,之后下降;干旱复水胁迫下,沙芥PcHSP26.5和PcHSP17.8基因的相对表达量均在干旱9 d达到顶峰,复水后下降。

    Abstract:

    On the basis of the joint analysis of transcriptome and proteome data, two small heat shock protein genes, PcHSP26.5 and PcHSP17.8, were cloned by RT-PCR using seedling leaves of Pugionium cornutum (L.) Gaertn., and analyzed by bioinformatics software. Nucleotides and encoded proteins were analyzed by Real-time PCR for the expression patterns of the 2 genes under high temperature, low temperature, NaCl stress, ABA stress and drought and rewatering stress. The open reading frames of PcHSP26.5 and PcHSP17.8 are 699 bp and 462 bp, respectively, encoding 282 and 153 amino acids, respectively, of which PcHSP26.5 contains an intron. The relative molecular masses of PcHSP26.5 and PcHSP17.8 are 26 480 and 17 490, respectively, and their theoretical isoelectric points are 6.36 and 5.99, respectively. They are hydrophilic proteins without transmembrane structure and signal peptide sites. Subcellular location prediction and phylogenetic tree analysis showed that PcHSP26.5 is mainly located in the mitochondria and belongs to the MII subfamily; while PcHSP17.8 is mainly located in the cytoplasm and belongs to the CI subfamily. The results of real-time fluorescent PCR showed that under high temperature stress, relative expression levels of PcHSP26.5 and PcHSP17.8 genes in Pugionium cornutum (L.) Gaertn. decreased after reaching the peak at 1 h; under low temperature stress, the relative expression level of PcHSP26.5 gene decreased after reaching the peak at 0.5 h, and the relative expression level of PcHSP17.8 gene was lower than the control within 0.5-12 h, and showed an upward trend at 24 h; under NaCl stress, relative expression levels of PcHSP26.5 and PcHSP17.8 genes decreased after reaching their peaks at 1 h and 3 h, respectively; under ABA stress, relative expression levels of the PcHSP26.5 and PcHSP17.8 decreased after reaching the peak at 2 h; under drought and rewatering stress, the relative expression levels of PcHSP26.5 and PcHSP17.8 genes reached their peak at 9 days of drought, and then declined after rewatering.

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于晓婧,王萍,武兆昕,杨永升,许珂,尚鹏.沙芥幼苗叶片小热激蛋白基因的克隆与表达[J].湖南农业大学学报:自然科学版,2021,47(6):.

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  • 在线发布日期: 2021-12-28
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