甘菊ClHSP70ClHSP90基因的克隆及表达分析
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北京市农林科学院科技创新能力建设专项(KJCX20140109)


Cloning and Expression of ClHSP70 and ClHSP90 Gene from Chrysanthemum lavandulifolium
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    摘要:

    该研究以甘菊(Chrysanthemum lavandulifolium)为实验材料,通过RTPCR方法从甘菊转录组数据中分离出热激蛋白合成相关基因,命名为ClHSP70ClHSP90。序列分析表明,ClHSP70基因ORF全长为2 559 bp,编码852个氨基酸,蛋白功能区预测表明含有典型的HSP70蛋白NBD和SBD保守结构域;ClHSP90基因ORF全长为2 094 bp,编码697个氨基酸,含有HATPase结构域和HSP90保守结构域。生物信息学分析表明,甘菊ClHSP70与大豆(Glycine max)和烟草(Nicotiana tomentosiformis)HSP70蛋白有较高的一致性, ClHSP90基因编码的氨基酸序列与紫茎泽兰(Ageratina adenophora)HSP90高度相似;实时荧光定量表达分析表明,在42 ℃处理不同时间,甘菊叶片中ClHSP70ClHSP90基因表达均在0.5 h时显著增加,1 h达到最大值,2 h后缓慢下降;不同组织表达分析表明,甘菊在42 ℃处理1 h后,ClHSP70在成熟叶中的表达量显著高于嫩叶和根等其他组织;ClHSP90在成熟茎中的表达量最高。研究说明,ClHSP70ClHSP90基因具有热激蛋白特征,参与了甘菊热胁迫应答过程,该研究结果为以后深入研究其基因功能奠定了基础。

    Abstract:

    Based on Chrysanthemum lavandulifolium RNAseq data, we obtained two code DNA sequences named ClHSP70 and ClHSP90 using RTPCR. Sequence analysis showed the predicted open reading frame (ORF) of ClHSP70 is 2 559 bp, encoding a protein of 852 amino acids. The conserver domain search exhibited that Nteminar contains typical NBD domain which belongs to HSP70 family. The predicted ORF of ClHSP90 is 2 094 bp, encoding a protein of 697 amino acids. The conserver domain search showed that Nteminar contains ATPasesuperfamily and HSP90 superfamily conservative structure. Bioinformatics showed that the amino acid sequences of ClHSP70 had high consistency with Glycine max and Nicotiana tomentosiformis HSP70, while ClHSP90 had high consistency with Ageratina adenophora HSP90. Realtime PCR demonstrated the expression patterns of ClHSP70 and ClHSP90 in leaves at 42 ℃. The expression of these two genes were both significantly upregulated after 0.5 h, reached the highest level after 1 h. Then the expression of both two genes decreased after 2 h, and remained at a relatively lower level after 4 h. In different plant tissues (root, stem, leaf) after 1 h at 42 ℃, ClHSP70 was highly expressed in mature leaves, lower in young, leaves, root, mature stems, young stems, and ClHSP90 was the highest in mature stems. The results showed that ClHSP70 and ClHSP90 participated the process of heat tolerance in Chrysanthemum lavandulifolium and laid a foundation for analyzing ClHSP70 and ClHSP90 gene function.

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罗 昌,陈东亮,程 曦,等.甘菊ClHSP70ClHSP90基因的克隆及表达分析[J].西北植物学报,2016,36(7):1321-1330

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  • 在线发布日期: 2016-08-16
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