马铃薯WOX基因家族的鉴定及在离体再生和非生物胁迫中的表达分析

Genome-wide Identification of Potato WOX Gene Family and Its Expression Analysis in in vitro Regeneration and Abiotic Stress

【目的】WUSCHE-相关同源盒(WUSCHEL-related homeobox,WOX)基因家族是植物特有的转录因子家族,在植物生长发育、干细胞分化调控、逆境胁迫响应等过程中扮演重要角色。开展马铃薯WOX基因家族鉴定与功能研究,将为马铃薯遗传改良提供优良基因资源与理论依据。【方法】基于拟南芥、番茄、烟草和水稻WOX蛋白序列,利用HMMER 3.0和BLASTP鉴定马铃薯WOX基因家族成员,使用MCScanX软件分析WOX基因家族成员在马铃薯种内及种间的共线性,并采用邻接法构建系统发育进化树。利用ExPASy、GSDS等软件分析马铃薯WOX基因家族成员理化性质、基因结构、蛋白motif、启动子区域转录因子结合位点。基于PGSC数据库中马铃薯转录组数据,分析StWOXs在不同组织和非生物胁迫下的表达模式;以可能参与离体再生过程的StWOX5作为候选基因,利用实时荧光定量PCR技术分析该基因在具有不同离体再生能力的4个马铃薯品种(系)再生过程中的表达情况。【结果】鉴定得到11个马铃薯WOX基因家族成员,分布在5条染色体上,分为WUS、中间和古老共3个进化分支,不同分支中StWOXs基因结构、蛋白motif组成、转录因子结合位点类型和数量存在不同程度的差异。表达模式结果表明,马铃薯WOX基因家族成员参与离体再生和非生物胁迫响应,StWOX4/5/11/13在愈伤组织中特异表达,StWUS、StWOX1/3c/4/5/13在甘露醇处理下上调表达,StWOX3a/11在NaCl处理下上调表达,StWOX4/5/13在热胁迫中上调表达,且StWOX5相对表达量与愈伤组织分化率呈正相关。【结论】不同马铃薯WOX基因具有潜在的功能多样性,StWOX5具有促进马铃薯离体再生分化的作用,参与了非生物胁迫反应。

【Objective】The WUSCHEL-related homeobox(WOX)gene family is plant-specific,which plays important roles in plant growth and development,regulation of stem cell differentiation,and response to adverse stress.Identification of potato WOX gene family may provide a theoretical basis and gene resources for potato genetic improvement.【Method】Based on the WOX protein sequences of Arabidopsis thaliana,tomato,tobacco and rice,all members of potato WOX gene family were identified by HMMER 3.0 and BLASTP,MCScanX was used for collinear analysis,and the phylogenetic tree was constructed using neighbor-joining method.Physical and chemical properties,gene structure,protein motif,and transcription factor binding sites of potato WOX members were analyzed by ExPASy,GSDS and other software.Expression patterns of StWOXs were analyzed based on potato transcriptome data from PGSC database.RT-qPCR assay was carried out to analyze relative expression of StWOX5 in four potato varieties(lines)with different in vitro regeneration abilities.【Result】11 members of potato WOX gene family were identified,distributed on five chromosomes,and classified into three clades,namely WUS,intermediate and ancient,with differences in gene structure,motif composition,transcription factor binding sites.The results showed that members of potato WOX gene family were involved in in vitro regeneration and abiotic stress responses.StWOX4/5/11/13 were specifically expressed in callus,StWUS and StWOX1/3c/4/5/13 were up-regulated under mannitol treatment,StWOX3a/11 were up-regulated under salt treatment,and StWOX4/5/13 were up-regulated under heat stress.The relative expression of StWOX5 was positively correlated with the differentiation rate.【Conclusion】Different potato WOX genes had potential functional diversity.StWOX5 may promote potato regeneration in vitro and participate in abiotic stress responses.