大豆GmNAC011基因的分离及对异黄酮合酶基因GmIFS2的调控分析

Isolation and Regulation Analysis of GmNAC011 Gene to Isoflavone Synthase Gene GmIFS2

通过对不同异黄酮含量品种大豆的转录组测序分析,发现GmNAC011基因在不同品种间的表达量存在显著差异。为了研究该基因的功能,根据大豆基因组序列设计引物,从大豆根系中克隆到GmNAC011基因完整的开放性阅读框(ORF)(Opening reading frame)序列,通过生物信息学对该序列进行分析,结果显示:该基因编码268个氨基酸,蛋白质分子量为31 k Da,理论等电点为8.3。进化树分析发现该蛋白与GmNAC2亲缘关系较近,属于NAC转录因子ATAF1亚家族。通过RT-PCR的方法分析了GmNAC011基因与异黄酮合酶基因GmIFS2在不同组织中的表达模式,结果显示GmNAC011与GmIFS2呈现相似的表达趋势,均为在花中表达量相对较低,而在根、茎、叶和荚中的表达量较高;通过酵母单杂交实验发现GmNAC011可以与GmIFS2基因启动子中的"CGTG"基序结合,这些结果说明了GmNAC011基因参与调控GmIFS2基因的表达,进而可以调控异黄酮的合成。

Based on the previous results of transcriptome sequencing,the expression of GmNAC011 was different in soybean cultivars that differed in isoflavonoid accumulation,thus we selected this gene for further analysis.Primers were designed according to the soybean genome sequences. The whole opening reading frame of GmNAC011 was isolated from soybean roots,encoding 268 amino acids with a calculated molecular mass of 31 kDa and a theoretical p I of 8. 3. Phylogenetic analysis showed that GmNAC011 was close to GmNAC2,belonging to ATAF1-like NAC transcription factor subgroup. We investigated the expression pattern of GmNAC011 and GmIFS2 in various tissues（ root,stem,leaf,flower and pod）,the expression model of GmNAC011 was similar with GmIFS2. The transcript levels of GmNAC011 and GmIFS2 were found to be higher in roots,stems,leaves and pods than flowers. And we found that GmNAC011 could binding to the motif ＂ CGTG＂ in the promoter of GmIFS2 by yeast one-hybrid. All results showed that GmNAC011 participate in regulating the expression of GmIFS2,subsequently the synthesis of isoflavonoid.