橡胶树感白粉病基因LFG1和LFG2的克隆及表达分析

Gene Cloning and Expression Analysis of LFG1 and LFG2 of Rubber Tree

白粉病是橡胶树的主要病害之一,为阐明橡胶树与白粉菌分子互作机制,本研究基于已报道的负调控基因LFG为基础,克隆出橡胶树中同源基因LFG1和LFG2,采用同源克隆方法,以橡胶树基因组DNA和cDNA为模板,PCR扩增HbLFG1和HbLFG2基因;利用生物信息学方法分析该基因及其同源蛋白;并分别对白粉菌侵染不同时间和不同病害等级的橡胶树叶片进行RT-PCR测定,检测HbLFG1和HbLFG2的基因表达量。HbLFG1和HbLFG2基因的核酸和氨基酸分析结果分别为:cDNA序列全长均为729 bp及由242个氨基酸编码组成,蛋白的分子量为27.220 kD和27.248 kD,等电点为8.24和8.63,正负电荷残基数为15和12与13和11,脂肪系数是132.52和130.50,不稳定系数为24.19和26.51,总平均亲水性0.958和1.011。两个基因都有7个跨膜结构域,不含有信号肽,蛋白都属于BI-1家族,蛋白模序有5处差异,两个基因与辣椒和柑橘的BI1-like蛋白同源性最为接近。HbLFG1随白粉菌侵染时间和病害等级的增加均呈现先上升后下降的趋势;HbLFG2随病害等级的上升呈现先上升后下降的趋势,但是随白粉菌侵染时间的增加呈现持续上升的趋势。HbLFG1和HbLFG2两个基因的表达量都与橡胶树白粉菌的侵染有关,但两者存在一定差异。本研究有助于深入阐释橡胶树与白粉菌分子互作机制。

Oidium heveae is one of the main diseases of rubber trees.To clarify the interaction mechanism of the rubber tree and the Oidium heveae.It based on the research of the reported negative regulation gene LFG.The study cloned the homologous genes LFG1 and LFG2 in rubber trees,which laid a foundation for elucidating the molecular interaction mechanism between rubber trees and powdery mildew.HbLFG1 and HbLFG2 genes were amplified by homologous cloning method using genomic DNA and eDNA of rubber tree as template.The gene and its homologous protein were analyzed by bioinformaties method.The rubber tree leaves of different disease grades were subjected to RT-PCR to detect the gene expression levels of HbLFG1 and HbLFG2.The results of nucleic acid and amino acid analysis of HbLFG1 and HbLFG2 genes were:The full length of the eDNA sequence was 729 bp and 242 amino acids.The molecular weight of the protein was 27.220 kD and 27.248 kD.The isoelectrie point was 8.24 and 8.63,the number of positive and negative charge residues is 15 and 12 and 13 and 11,the fat coefficient is 132.52 and 130.50,the instability coefficient is 24.19 and 26.51,the total average hydrophilicity of 0.958 and 1.011.Both genes have seven transmembrane domains,do not contain signal peptides,proteins belong to the BI-1 family,and there are five differences in protein motifs.The two genes are closest to the BIl-like proteins of pepper and citrus.HbLFG1 increased first and then decreased with the increase of infection time and disease grade.HbLFG2 showed a trend of increasing first and then decreasing with the increase of disease grade, but it continued to increase with the increase of infection time of powdery mildew.The expression levels of both HbLFG1 and HbLFG2 genes are related to the infection of powdery mildew of rubber tree,but there are some differences between them.These will help to explain the molecular interaction mechanism between rubber tree and powdery mildew.