甜菊耐寒相关基因SrDREB1As的克隆与转录因子特性分析

Molecular Cloning and Transcription Factor Characterization of Cold Tolerance Related Genes-SrDREB1As in Stevia rebaudiana Bertoni

AP2/ERF转录因子家族中DREB亚族的A-1亚组基因对植物耐寒性具有显著的调控作用。本研究通过同源克隆的方法在甜菊中获得2个A-1亚组基因,分别命名为SrDREB1A1和SrDREB1A2(NCBI基因登录号分别为MK163640和MK163641)。SrDREB1A1 ORF全长660 bp,编码219个氨基酸残基,预测的蛋白分子量为24.58 kDa,等电点为5.42。SrDREB1A2 ORF全长630 bp,编码209个氨基酸残基,预测的蛋白分子量和等电点分别为23.21 kDa和5.41。进化树分析发现,SrDREB1A1和SrDREB1A2分别与紫茎泽兰的AaCBF及菊花的DgDREB1A亲缘关系最近,序列分析发现二者都具有一个典型的AP2结构域。进一步对蛋白功能研究发现,SrDREB1A1和SrDREB1A2都定位在细胞核并且具有转录激活活性。本研究将通过转基因手段为提高甜菊的耐寒性提供重要的基因资源。

The DREB(dehydration responsive element binding protein)subfamily A-1 group genes of plant AP2/ERF transcription factor family play a significant role in regulating cold tolerance of plant.In this study,we isolated two A-1 group genes by homologous cloning,and designated them as SrDREB1A1 and SrDREB1A2(NCBI accession number MK163640 and MK163641).The open reading frame(ORF)of SrDREB1A1 was 660 bp and encoded protein of 219 amino acid residues with predicted molecular weight of 24.58 kDa and isoelectric point(PI)of 5.41.The cDNA of SrDREB1A2 was 630 bp and the encoding product comprised 209 amino acid residues with predicted molecular weight and PI of 23.21 kDa and 5.41,respectively.The phylogenetic analysis revealed that SrDREB1A1 and SrDREB1A2 had the closest relationship with AaCBF of Ageratina adenophora and DgDREB1A of chrysanthemum.Meanwhile,both SrDREB1A1 and SrDREB1A2 possessed the conserved AP2 domain.Further functional analysis showed that SrDREB1A1 and SrDREB1A2 extensively located in the nuclear and exhibited strong transactivation activities.Our study provides important gene resources for improving cold tolerance of stevia by transgenic manipulation in the future.