白菜叶形发育相关基因BrLOM2的克隆与表达分析

Cloning and Expression Analysis of BrLOM2,a Gene Involved in Lobed Leaf Development in Pakchoi

为了揭示白菜(Brassica rapa ssp.chinensis)BrLOM2基因在叶发育中的调控机制,以白菜圆叶、裂叶近等基因系为材料,同源克隆得到BrLOM2基因,分析其时空表达特性及对激素的响应。结果显示,BrLOM2的ORF全长为549 bp,编码182个氨基酸,含有GRAS家族保守结构域,定位于细胞核,两种材料中的氨基酸序列在保守结构域存在两处突变。进化分析表明,BrLOM2属于十字花科LOM2分支,与大白菜(XP_009138823.1)亲缘关系最近。RT-PCR结果显示,BrLOM2在圆叶白菜中的表达量高于同期裂叶白菜,两种材料茎尖、第1片叶中表达量最高;裂叶中的表达量与叶缘裂刻数呈现同步增加的趋势,6-BA和GA_3分别抑制和促进叶片中的表达,说明BrLOM2低表达可能是裂叶表型产生的主要原因。

To elucidate the mechanism underlying development of leaf marginal lobes in pakchoi,an LOM2 homologous gene,Br LOM2,was isolated respectively from near-isogenic lines with and without lobed leaves in pakchoi using the homology-based cloning methods. The Br LOM2 gene contains an open reading frame of 549 bp in length encoding a protein of 182 amino acids. Sequence analysis showed that the Br LOM2 protein contained conservative structure domains of the GRAS family. The BrLOM2 sequence from genomic DNA showed no intron and phylogenetic analysis suggested that BrLOM2 shares high homology with known LOMs from other plant species. Moreover,the putative Br LOM2 protein was closely clustered with several Brassica LOMs. q RT-PCR analysis revealed that Br LOM2 gene has higher expression in leaves and shoot apexes,and lower expression in flowers. Further more,BrLOM2 gene has higher expression level in pakchoi lines with smooth leaves than in the lines with lobed leaves. The expressionof BrLOM2 in pakchoi lines with lobed leaves became higher,while the number of lobes in them had the synchronized trend. In leaves,the expression of BrLOM2 was affected by application of exogenous GA3 and 6-BA hormone treatments. Interestingly,GA3 treatment increased the BrLOM2 gene expression,whereas 6-BA treatment decreased it. In summary,the above-mentioned results indicated that the reduced transcriptional expression of BrLOM2 gene was the major factor contributing to leaf marginal lobes.