龙眼生长素受体基因TIR1的克隆及表达分析

为进一步明确龙眼生长素受体基因TIR1的结构和功能,本试验采用RT-PCR和RACE-PCR对龙眼胚性愈伤组织2个TIR1基因(分别命名Dl TIR1-1和Dl TIR1-2)进行克隆,并进行生物信息学和表达分析。研究结果表明:Dl TIR1-1全长4 343 bp,包含ORF 1 755 bp,编码584个氨基酸(Gen Bank登录号:KR558759),而Dl TIR1-2全长2 821 bp,包含ORF 1 926 bp,编码641个氨基酸(Gen Bank登录号:KR558760)。生物信息学分析结果表明,Dl TIR1-1和Dl TIR1-2理化性质较为一致,均属于不稳定蛋白,不含信号肽,具有跨膜螺旋;亚细胞定位于细胞质中,分别含有31个和45个蛋白磷酸化位点;系统进化树分析显示,Dl TIR1-1与十字花科的亚麻荠和甜菜处于同一分支,而Dl TIR1-2与甜橙和胡杨等木本植物保持较近的遗传距离。q PCR结果表明,Dl TIR1-1和Dl TIR1-2在龙眼体胚发生过程中和不同组织器官中的表达模式较为一致,且在非胚性愈伤组织、根和花中表达量最高;此外,在一定的浓度范围内,IAA、ETH、ABA和GA3均能适当提高龙眼TIR1的表达量,而添加Me JA却明显抑制TIR1的转录。上述结果表明,Dl TIR1-1和Dl TIR1-2可能具有相似功能,且均参与龙眼非胚性愈伤组织形成、根系分化以及花器官发育,此外龙眼生长素信号转导途径可能受多种植物激素调控。

Cloning and Expression Analysis of Auxin Receptor GeneTIR1 from Dimocarpus longan Lour.

To investigate the characteristic and function of TIR1 in longan, RT-PCR and RACE-PCR were used to clone 2 TIR1 genes(named DlTIR1-1 and DlTIR1-2)from embryogenic callus of Dimocarpus longan Lour., and then the expression patterns of the 2 genes were analyzed by qPCR. The results showed that, the sequence of DlTIR1-1 was 4 343 bp, containing ORF sequence 1 755 bp, encoding 584 amino acids(GenBank accession number KR558759), and the sequence of DlTIR1-2 was 2 821 bp, containing ORF 1 926 bp, encoding 641 amino acids(GenBank accession number KR558760). Bioinformatics analysis showed that the physical and chemical properties of DlTIR1-1 and DlTIR1-2 were similar, they were unstable proteins, had no signal peptide, but had transmembrane structure. Phylogenetic tree analysis indicated that DlTIR1-1 belonged to the same branch with Camelina sativa and Beta vulgaris subsp, when DlTIR1-2 kept close genetic distance with Populus euphratica and Citrus sinensis. qPCR results showed that DlTIR1-1 and DlTIR1-2 had the consistent expression pattern in somatic embryogenesis, different tissues and treatments by different hormones, and both genes had the highest expressions in non-embryogenic callus, roots and flowers of longan. In addition, DlTIR1-1 and DlTIR1-2 had different expression patterns under different treatments of IAA, ABA, GA3, MeJA and ETH. The results indicated that, DlTIR1-1 and DlTIR1-2 had the similar function in the process of NEC growth, root differentiation and floral organ development of longan. In addition, the auxin signal transduction of longan might be regulated by several kinds of hormones.