甘蓝型油菜BnEXO基因家族功能与表达分析

Gene Family Function and Expression Analysis of BnEXO in Brassica napus

EXORDIUM(EXO)基因在拟南芥中被确定为油菜素内酯(BR)响应基因,通过介导细胞扩张促进植物生长。为探究EXO基因在甘蓝型油菜中的功能以及在花期不同组织中的表达规律,以甘蓝型油菜中双6号为材料,克隆EXO基因的编码区序列命名为BnEXO,并进行生物信息学分析,利用实时荧光定量技术对BnEXO基因在甘蓝型油菜花期根、茎、叶、花瓣、花蕾和角果皮中的相对表达量进行测定。结果表明,BnEXO基因的CDS序列为945 bp,BnEXO为稳定亲水性非跨膜蛋白,属于分泌蛋白,在细胞外表达,蛋白的二级结构以无规则卷曲为主。不同组织中的表达分析结果表明,BnEXO基因在不同组织中的表达量由高到低依次为花瓣、角果皮、花蕾、茎、根、叶,在花瓣中的表达量最高。此外,以拟南芥中8个EXO基因家族成员的蛋白序列为基础,鉴定到20个甘蓝型油菜EXO基因(BnaEXO),11个白菜EXO基因(BraEXO)和11个甘蓝EXO基因(BoEXO)。大多数基因家族成员编码蛋白质为稳定性蛋白,定位在细胞外,长度为271~411个氨基酸,等电点预测为5.76~9.60,分子质量为28.76~46.21 ku。系统发育分析将EXO基因分为EXOA、EXOB、EXOC、EXOD和EXOE共5个亚组,EXOB亚组中成员最少。基因结构分析表明,大多数成员只含有1个外显子,没有内含子,EXO基因家族成员序列具有高度的保守性。甘蓝型油菜中成员的启动子区域顺式元件分析结果表明,BnaEXO基因在植物的生长发育及逆境胁迫中发挥重要作用。

The EXORDIUM(EXO)gene was identified in Arabidopsis thaliana as an brassinosteroid(BR)-responsive gene that promotes plant growth by mediating cell expansion.In order to investigate the function of EXO gene in Brassica napus and its expression pattern in different tissues at flowering stage,we used B.napus Zhongshuang 6 as the material,cloned the sequence of the coding region of the EXO gene named BnEXO,and carried out bioinformatics analysis,and used Real-time Fluorescence Quantification to determine the relative expression of BnEXO gene in B.napus in roots,stems,leaves,petals,buds,and pericarps at flowering stage.The results showed that the CDS sequence of BnEXO gene was 945 bp,BnEXO was a stable hydrophilic non-transmembrane protein,which belonged to secreted proteins and was expressed extracellularly,and the secondary structure of the protein was dominated by the random coil.The results of expression analysis in different tissues showed that the expression of BnEXO gene in different tissues was in the order of petals,pericarps,buds,stems,roots and leaves,and the highest expression was found in petals.In addition,20 B.napus EXO genes(BnaEXO),11 B.rapa EXO genes(BraEXO),and 11 B.oleracea EXO genes(BoEXO)were identified in this study based on the protein sequences of eight EXO gene family members in A.thaliana.Most proteins of gene family members were stable proteins,localized extracellularly,with amino acid lengths ranging from 271 to 411 aa,isoelectric point predictions ranging from 5.76 to 9.60,and molecular masses ranging from 28.76 to 46.21 ku.Phylogenetic analysis classified the EXO genes into five subgroups,EXOA,EXOB,EXOC,EXOD,and EXOE,with the least number of members in the EXOB subgroup.Gene structure analyses showed that most members contained only one exon and no intron,and the sequences of EXO gene family members were highly conserved.The results of cis-element analysis of the promoter region of the members in B.napus indicated that the BnaEXO genes play important roles in plant growth and development and in adversity stress.