野生番茄SpIDD1基因的表达分析及其VIGS载体构建

Expression Analysis and VIGS Vector Construction of the SpIDD1 Gene in Wild Tomato

为鉴定野生番茄的SpIDD1基因并探究其功能,以野生醋栗番茄‘PI365967’为试验材料,利用RT-PCR克隆得到SpIDD1基因的CDS序列,全长1 020 bp,编码339个氨基酸,与普通番茄参考基因组序列相比,SpIDD1只存在一个核苷酸位点的差异。蛋白序列比对和结构分析表明,Sp IDD1含有1个核定位信号和4个保守锌指结构域(C2H2·C2H2·C2HC·C2HC),是一个典型的IDD蛋白。系统进化分析表明,SpIDD1与马铃薯和辣椒的IDD蛋白亲缘关系最近。荧光定量PCR显示,SpIDD1在叶、顶芽和果实中的表达较高,并受干旱胁迫的显著诱导。利用TRV-VIGS系统构建了Sp IDD1的基因沉默载体,并成功侵染番茄。研究结果表明,SpIDD1作为一个典型的IDD基因,很可能参与了番茄的干旱胁迫。本研究为进一步研究该基因的功能及作用机理提供理论依据。

To identify the SpIDDl gene and explore its function in wild tomato, this study took Solarium pimpinellifolium 'PI365967' as a test materials, cloned the CDS sequence of SpIDDl gene with 1 020 bp full length and encoded for 339 amino acids. There was only one nucleotide divergence compared with the reference genome sequence of the cultivated tomato Solanum lycopersicum. Sequence alignment and structure analysis demonstrated that SpIDDl contained a nuclear localization signal (NLS) and four conserved zinc finger domain (C2H2·C2H2· C2HC·C2HC). It was a typical IDD protein. Phylogeny analysis showed that SpIDDl might have the closest relationship with the IDD of potato and pepper. Quantitative real-time PCR analysis illustrated that SpIDDl expressed highest in leaf, terminal bud and fruit. It was significantly induced by drought stress. Using the Tobacco rattle virus (TRV)-based VIGS system, we constructed the pTRV2-Sp/DD/ vector and infected the cotyledon of Solarium pimpinellifolium 'PI365967' successfully. The results suggested that SpIDDl, as a typical IDD gene, might participate in the drought stress response in tomato. It might provid the foundation for further study the function of SpIDDl.