芜菁BrrHMA2.1和BrrHMA2.2基因的克隆与表达

摘要

该研究以芜菁(Brassica rapa var.rapa)为材料,克隆得到重金属ATP酶(HMA)家族1对同源基因BrrHMA2.1(GenBank登录号:MG_283237)和BrrHMA2.2(GenBank登录号:MG_283238),并对其蛋白质序列特征和基因表达模式进行分析.结果表明:(1) BrrHMA2.1和BrrHMA2.2基因的全长开放阅读框分别为2 619和2 724 bp,分别编码872和907个氨基酸;序列结构分析显示,BrrHMA2.1和BrrHMA2.2蛋白含有6个跨膜区和HMA蛋白家族保守结构域;系统进化树结果显示,BrrHMA2.1和BrrHMA2.2蛋白与拟南芥HMA家族成员AtHMA2进化关系最近.(2)亚细胞定位结果表明,BrrHMA2.1和BrrHMA2.2蛋白都定位于细胞膜上.(3)qRT-PCR分析表明,芜菁生长初期BrrHMA2.1和BrrHMA2.2基因在叶中的表达量最高;随着生长时间的延长,叶中的表达量逐渐降低,而根中的表达量逐渐增加.(4)研究发现,BrrHMA2.1受Cd2+、Zn2+、Na+、Mg2+胁迫诱导表达,BrrHMA2.2受Cd2+、Na+、Cu2+胁迫诱导表达,表明2个基因可能参与这些金属离子的转运过程.该研究结果为进一步研究植物HMA基因在重金属吸收和转运过程中的功能奠定了基础.%In the present study,a pair of homologous genes BrrHMA2.1 (GenBank is MG_283237) and BrrHMA2.2 (GenBank is MG_283238) of the heavy metal ATPase (HMA) family in turnip (Brassica rapa var.rapa) was cloned and their protein sequence characteristics and gene expression patterns were analyzed.(1) The full-length open reading frames of these two genes were 2 619 and 2 724 bp and they encoded 872 and 907 amino acids,respectively.Sequence structure analysis showed that both BrrHMA2.1 and BrrHMA2.2 proteins included six transmembrane regions and conserved domains of HMA family.Phylogenetic tree analysis showed that BrrHMA2.1 and BrrHMA2.2 proteins had the closest evolutionary homology with AtHMA2.(2) The results of subcellular localization indicated that both BrrHMA2.1 and BrrHMA2.2 proteins were localized in the plasma membrane.(3) qRT-PCR analysis showed that the expression levels of BrrHMA2.1 and BrrHMA2.2 genes were much higher in leaves at the early stage of turnip.However,the expression levels of these two genes in leaves gradually decreased whereas those in roots gradually increased with the increasing growth time.In addition,the results also found that BrrHMA2.1 gene was induced by Cd2+,Zn2+,Na+ and Mg2+ while BrrHMA2.2 gene was induced by Cd2+,Na+ and Cu2+ stresses,indicating these two genes may play roles in coping with these metal ions.This study provides foundation for further functional studies on plant HMA genes in heavy metal tolerance or transport.