烟草氧化胁迫相关基因NtOSA1的克隆表达及亚细胞定位分析
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国家自然科学基金(31500205);


Cloning, Expression Characterization and Subcellular Localization of NtOSA1 Gene from Nicotiana tabacum
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    摘要:

    植物类蛋白激酶Abc1家族(activity of bc1 complex)在植物生长发育及响应非生物胁迫中起着重要的作用。该研究通过将拟南芥AtOSA1蛋白氨基酸序列与烟草转录组数据进行比对,利用巢式PCR技术克隆得到1个烟草氧化胁迫相关Abc1家族基因NtOSA1,NtOSA1与拟南芥AtOSA1基因具有72.89%的一致性。NtOSA1基因开放阅读框长度为2 283 bp,编码760个氨基酸,含有典型的ABC1结构域、一个激酶结构域、叶绿体定位信号肽和两个跨膜结构域。采用实时荧光定量PCR技术,对NtOSA1基因在烟草不同组织以及氧化胁迫、盐胁迫等处理下的表达分析表明:NtOSA1基因的表达具有组织特异性,主要在叶片中表达;NtOSA1基因在H2O2和NaCl处理后表达量上升,均在处理后6 h达到最大值,分别为处理前的1.95和2.69倍。亚细胞定位结果表明,NtOSA1蛋白定位在细胞叶绿体上,与预测结果一致。研究表明,NtOSA1基因参与了烟草抗氧化胁迫和盐胁迫的响应。

    Abstract:

    Members of the activity of bc1 complex (Abc1) family are protein kinases that are functionally diverse proteins with multiple roles in the regulation of plant growth and development and abiotic stress tolerance. According to the amino acid sequence of AtOSA1 and the transcriptome data of Nicotiana tabacum, a Abc1 like gene was isolated and designated as NtOSA1, as for the high identity with the AtOSA1 (Arabidopsis thaliana oxidative stressrelated Abc1like protein). The open reading frame (ORF) of the NtOSA1 gene is 2 283 bp which encoded a deduced protein including 760 amino acid residues. The protein sequence of NtOSA1 possesses a conserved ABC1 domain, one kinase domain, one chloroplast localization signal peptide and two transmembrane spans. The relative expression of NtOSA1 was determined by quantitative realtime PCR (qRTPCR). The results showed that NtOSA1 gene was expressed particularly in leaves, but also in flowers. After treatment of H2O2 and NaCl for 6 h, the expression of NtOSA1 increased to the maximum, which was 1.95 and 2.69 folds of the control, respectively. When fused to green fluorescent protein (GFP), NtOSA1 localized to the chloroplast in tobacco protoplasts, which was consistent with the prediction results of the software TargetP 1.1 Server. These results suggested that NtOSA1 may be involved in response to oxidative and salt stress in tobacco.

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刘继恺,高永峰,吴婵娟,等.烟草氧化胁迫相关基因NtOSA1的克隆表达及亚细胞定位分析[J].西北植物学报,2016,36(8):1499-1506

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  • 在线发布日期: 2016-08-30
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