甘蓝纤维素合成酶基因BoCesA的克隆与序列分析  

Cloning and sequence analysis of the BoCesAgene in Brassica oleracea L.

作　　者：徐凤凤[1] 向娟[1] 李双桃[1] 石锦[1] 李殿波[1] 连蔚然 赵冰[1] 郭仰东[1] 

机构地区：[1]中国农业大学农学与生物技术学院/设施蔬菜生长发育调控北京市重点实验室,北京100193

出　　处：《中国农业大学学报》2015年第6期131-137,共7页Journal of China Agricultural University

基　　金：国家重点基础研究发展计划(2012CB113900);北京市现代农业产业体系(BLVT-03)

摘　　要：为研究甘蓝纤维素合成酶BoCesA基因在干旱胁迫下的表达模式,通过同源克隆得到甘蓝纤维素合成酶基因的cDNA全长,利用生物信息学软件分析该基因的特征;构建含BoCesA和GFP的融合表达载体,通过基因枪转化洋葱表皮细胞试验对该基因进行亚细胞定位;利用荧光定量PCR分析基因在不同组织中的表达情况及干旱胁迫下的表达模式。甘蓝纤维素合成酶BoCesA基因的cDNA全长为2 721bp,编码906个氨基酸,在N端含有锌指结构域和2个跨膜区,C端含有6个跨膜区,除包含植物纤维素合成酶基因共有的保守区外,还包含2个高突变区。通过氨基酸序列比对分析,发现甘蓝的该基因与拟南芥的AtCesA3基因同源性较高。亚细胞定位表明BoCesA基因初步定位于细胞质膜上。荧光定量PCR分析表明该基因在叶中的表达量高于茎和根中的表达量,在NaCl、PEG处理下BoCesA表达量呈现先升高后下降的趋势。通过对甘蓝纤维素合成酶BoCesA基因在NaCl、PEG胁迫下的表达分析,预测该基因对干旱胁迫有响应,它的表达受干旱胁迫的影响,这为进一步研究基因功能奠定了基础。To study the mechanism of abiotic stress resistance,expression variation of BoCesAgene was studied after drought stress treatment.Homology-based cloning was used to amplify the gene.Sequence structure was analyzed by using bioinformatics softwares.To investigate the intracellular localization of the protein,GFP sequence was fused downstream to the BoCesA gene and the fusion gene BoCesA：：GFP was transferred into onion epidermal cells by biolistic method.Real-time PCR analysis revealed that the expression of BoCesA gene was significantly increase under stress.The coding region of the gene is 2 721 bp,encoding 906 amino acids.And it contains Zinc-binding domain,high specific areas and eight transmembrane domains.Subcellular location analysis indicates that the BoCesA gene is located in the cytoplasmic membrane.Real-time PCR analysis showed that the amount of gene expression in leaves is higher than those in stems and roots.The expression of BoCesA gene responds to and is affected by drought stress.

关 键 词：甘蓝 纤维素合成酶基因 亚细胞定位 组织特异性表达 荧光定量PCR 

分 类 号：S635[农业科学—蔬菜学]