大麦VQ基因家族鉴定及表达分析

Identification and expression analysis of VQ gene family in barley

【目的】鉴定大麦VQ基因家族成员并进行表达分析,为大麦VQ基因的功能挖掘提供理论依据。【方法】从大麦基因组中鉴定VQ基因家族成员,利用生物信息学方法对其结构特征及编码蛋白序列进行分析,基于转录组测序数据及实时荧光定量PCR方法进行大麦组织表达模式、盐胁迫和生物胁迫分析。【结果】在大麦基因组中鉴定出29个HvVQ基因(HvVQ1~HvVQ29),HvVQ蛋白序列平均长度较短(214 aa),多数HvVQ蛋白为碱性或偏中性蛋白,HvVQ基因不均地分布在大麦染色体上,定位于细胞核中。29个HvVQ蛋白均含有保守基序FxxxVQxhTG,近90%的HvVQ基因不含内含子。进化分析将大麦、拟南芥与水稻的VQ基因家族成员分为7个亚族(Ⅰ~Ⅶ),HvVQs基因不均地分布在Ⅱ~Ⅶ亚族中。大麦与水稻的共线性基因对数(17对)远多于与拟南芥的共线性基因对数(1对),种内共线性分析发现1对共线性基因对,非同义替换率/同义替换率(Ka/Ks)计算发现HvVQ蛋白主要处于纯化选择状态。HvVQ基因启动区富含生长发育作用元件、非生物胁迫反应元件和激素反应元件,种类及分布均呈多样性。对蛋白网络预测分析推断其与HvWRKY的2类亚族(Ⅱ-c和Ⅲ)存在互作关系。大多数HvVQ基因在组织中表达,HvVQ19在受到盐胁迫时表达量明显上调,在根尖和根伸长区表达量分别上调1.40和1.10倍;对其中10个HvVQ基因进行实时荧光定量PCR检测,HvVQ2基因在蚜虫和黄矮病毒胁迫下表达量均显著下调(倍数变化<0.5为显著抑制,>2.0为显著诱导),HvVQ7和HvVQ15基因在蚜虫和黄矮病毒胁迫下表达量上调最显著,其他7个HvVQ基因也均表现出差异表达。【结论】HvVQ基因家族成员存在序列保守性和进化性,其表达具有组织特异性,大多数成员表达水平受盐胁迫调控,有大量的激素响应元件,推测在生物和非生物胁应答中发挥重要调控作用。

【Objective】In this study,gene family members of VQ in barley were identified expression analysis was conducted,which provided theoretical basis for functional mining of barley VQ gene.【Method】In this study,the VQ gene was identified from the barley,and its structural features and coding protein sequences were analyzed using a bioinforma-tics approach.Barley tissue expression patterns,salt stress and biotic stress were analyzed based on transcriptome sequen-cing data and real-time fluorescent quantitative PCR(qRT-PCR).【Result】A total of 29 HvVQ genes(HvVQ1-HvVQ29)were identified in the barley genome.The average length of HvVQ protein sequences was short(214 aa),most HvVQ pro-teins were basic or neutral,and HvVQ genes were unevenly distributed on the chromosomes of barley and localized in the nucleus.All 29 HvVQ proteins contained the conserved motif FxxxVQxhTG,and nearly 90%HvVQ genes have no in-trons.Phylogenetic analysis were classified the VQ family genes of barley,Arabidopsis and rice into 7 subfamilies(Ⅰ-Ⅶ),and HvVQs genes was unevenly distributed in subfamiliesⅡ-Ⅶ.Interspecies covariance analysis showed that the number of covariate gene pairs with rice(17 pairs)was much higher than that with Arabidopsis(1 pair),and intraspecies covariance analysis found 1 covariate gene pair.Nonsynonymous substitution rate/synonymous substitution rate(Ka/Ks)calculations revealed that HvVQ proteins were mainly in the purifying selection state.The promoter region of the HvVQ genes was rich in growth and developmental elements,abiotic stress-responsive elements,and hormone-responsive ele-ments,with a diversity of species and distribution.Predictive analysis of the protein network inferred that it interacted with two subfamilies of HvWRKYs(II-c and III).Most of the HvVQ genes were expressed in tissues,and HvVQ19 expression was up-regulated when subjected to salt stress,the expression levels in the root tip and root elongation regions were up-regulated by 1.40 and 1.10 times respectively.qRT-PCR was performed on 10 of the HvVQ genes,of which HvVQ2 gene was significantly down-regulated under both aphid and yellow dwarf virus stresses(fold change<0.5 for significant repression and>2.0 for significant induction),HvVQ7 and HvVQ15 genes were the most significantly up-regula-ted under aphid and yellow dwarf virus stresses,and the other 7 HvVQ genes all showed differential expression.【Conclu-sion】HvVQ gene family members are conserved and evolved in sequence,their expression is tissue specific,most mem-bers expression levels are regulated by salt stress,have a large number of hormone response elements,and are presumed to play important regulatory roles in biotic and abiotic stresses.