番茄SlNRT2基因家族鉴定、启动子克隆及表达分析

Identification and characterization of the SlNRT2 gene family and cloning and expression analysis of SlNRT2 in Solanum lycopersicum

硝酸盐转运蛋白2(Nitrate transporter 2,NRT2),是一种高亲和硝酸盐转运蛋白,在植物对硝酸盐的吸收和转运过程中发挥重要作用。本研究通过生物信息学方法对番茄(Solanum lycopersicum)NRT2(SlNRT2)家族基因进行鉴定,利用热图分析SlNRT2家族成员基因表达模式,并利用启动子嵌合GUS报告基因转化拟南芥对SlNRT2启动子活性进行分析。结果表明番茄基因组中鉴定到5个NRT2基因,定位于细胞膜,氨基酸序列长度介于458~531 aa,等电点介于9.05~9.33,不稳定性指数介于31.84~40.70。基因结构分析表明,该基因家族由3~4个外显子构成。染色体定位分析显示,SlNRT2基因分布于3条染色体上。系统进化和保守结构域分析显示,SlNRT2分别归属于I和II两个类群,具有10种保守基序,蛋白保守性很高。番茄不同组织热图分析表明SlNRT2.1和SlNRT2.2呈组成型表达;SlNRT2.3~SlNRT2.5主要在根部表达。启动子顺式作用元件及GUS组织化学染色分析结果表明,SlNRT2.4启动子驱动GUS基因在拟南芥根部特异性表达,而SlNRT2.3和SlNRT2.5除驱动GUS基因在拟南芥根部表达,叶片也有表达。本研究为后续开展SlNRT2基因的功能研究提供依据,同时为培育氮素高效利用型转基因番茄奠定了基础。

Nitrate transporter 2 is a high-affinity nitrate transporter,which plays important roles in nitrate uptake and transport in plants.In this study,we performed a comprehensive bioinformatic analysis of NRT2 gene family in tomato genome,and detected the tissue expression pattern of SlNRT2 by heat map analysis.Furthermore,we conducted SlNRT2 promoter activity analysis by using the promoter chimeric GUS reporter gene tansform Arabidopsis thaliana.The bioinformatic analysis showed that there were 5 Sl-NRT2s detected in the tomato genome,which mainly located in cell membrane,with protein length ranged from 458 to 531 aa,pI value distributed from 9.05 to 9.33 and instability index ranged from 31.84 to 40.70.Gene structure analysis showed that the number of exons within SlNRT2 genes was 3−4.Chromosome mapping analysis revealed that the 5 SlNRT2s were distributed on 3 chromosomes.The results of phylogenetic and conserved motif analysis showed that SlNRT2 gene family was divided into I and II two groups,it contained ten major conserved motifs,and the protein domain of SlNRT2 family is conserved.Heat map results demonstrated that SlNRT2.1and SlNRT2.2 were expressed in all the tissues,while SlNRT2.3−SlNRT2.5 were specifically expressed in tomato roots.Analysis of cis-acting elements in promoter and histochemical staining revealed that SlNRT2.4 promoter drove GUS gene to accumulate specifically in Arabidopsis roots,while SlNRT2.3 and SlNRT2.5 expressed in both roots and leaves in Arabidopsis thaliana,which was accordance with the results of heat map analysis.These results will provide a basis for further functional research of SlNRT2 protein,and also lay a foundation for cultivating transgenic tomato with high nitrogen utilization efficiency.