忽地笑ABCG5转运蛋白基因(LaABCG5)的克隆和表达分析

Cloning and expression analysis of ABCG5 transporter gene from Lycoris aurea(LaABCG5)

通过分析忽地笑〔Lycoris aurea(L'Hér.)Herb.〕比较转录组数据,采用cDNA末端快速扩增(RACE)技术克隆到1个ABC转运蛋白基因LaABCG5。LaABCG5基因全长1896 bp,编码631个氨基酸。LaABCG5的理论相对分子质量为70451,理论等电点为pI 8.37。LaABCG5的二级结构中,α-螺旋、延伸链、β-转角和无规则卷曲的氨基酸所占比例分别为48.81%、13.47%、5.07%和32.65%。聚类分析结果显示:LaABCG5与拟南芥〔Arabidopsis thaliana(Linn.)Heynh.〕ABCG5转运蛋白的亲缘关系最近。LaABCG5与海枣(Phoenix dactylifera Linn.)、油棕(Elaeis guineensis Jacq.)、番木瓜(Carica papaya Linn.)、毛果杨(Populus trichocarpa Torr.et Gray)和银白杨(Populus alba Linn.)的ABCG5转运蛋白的同源性在62%~67%之间。qRT-PCR检测结果显示:LaABCG5基因在忽地笑的根、鳞茎、叶片、花瓣和雌蕊中均有表达,其中,在叶片和雌蕊中的相对表达量显著高于其他组织;100μmol·L^-1 MeJA处理后6 h,忽地笑叶片中LaABCG5基因的相对表达量显著高于对照(DMSO处理后6 h)。跨膜结构域和亚细胞定位结果显示:LaABCG5定位于细胞膜。研究结果显示:LaABCG5基因的表达具有组织特异性,且受MeJA调控;LaABCG5的定位与其功能相吻合。

Based on analyzing comparative transcriptome data of Lycoris aurea(L'Hér.)Herb.,an ABC transporter gene LaABCG5 was cloned by rapid amplification of cDNA ends(RACE)technology.The full-length of LaABCG5 gene is 1896 bp,which encodes 631 amino acids.Theoretical relative molecular mass of LaABCG5 is 70451,and its theoretical isoelectric point is pI 8.37.In the secondary structure of LaABCG5,the percentage of amino acids ofα-helix,extended strand,β-turn,and random coil is 48.81%,13.47%,5.07%,and 32.65%,respectively.The result of cluster analysis shows that LaABCG5 is the closest to ABCG5 transporter from Arabidopsis thaliana(Linn.)Heynh.The homology of LaABCG5 with ABCG5 transporters from Phoenix dactylifera Linn.,Elaeis guineensis Jacq.,Carica papaya Linn.,Populus trichocarpa Torr.et Gray,and Populus alba Linn.is 62%-67%.The detection results of qRT-PCR show that LaABCG5 gene expresses in root,bulb,leaf,petal,and pistil of L.aurea,in which,its relative expression in leaf and pistil is significantly higher than that in other tissues;relative expression of LaABCG5 gene from leaf of L.aurea after 100μmol·L^-1 MeJA treatment for 6 h is significantly higher than that in the control(after DMSO treatment for 6 h).The results of transmembrane domain and subcellular localization show that LaABCG5 is localized on the cell membrane.It is suggested that the expression of LaABCG5 gene shows tissue specific and is regulated by MeJA,and the location of LaABCG5 is consistent with its function.