不结球白菜BcHSP70-1基因的克隆与进化及其表达分析

Clone,Phylogenetic and Expression Analysis of BcHSP70-1 Gene fromBrassica campestris ssp.chinensis

利用已报导的拟南芥和甘蓝型油菜同源基因序列设计克隆引物,从不结球白菜中克隆了BcHSP70-1基因。在BcHSP70-1基因测序后,对该基因编码蛋白质的理化性质、亚细胞定位和跨膜结构域等生物信息学分析结果表明,BcHSP70-1编码的蛋白质是亲水蛋白,定位在细胞质中,该基因ORF全长1 950bp,编码649个氨基酸,有1个内含子和2个外显子,内含子为324bp;对BcHSP70-1直系同源基因外显子和内含子比对分析显示,内含子的差异更显著。实时定量PCR表达分析表明,BcHSP70-1的表达量在不同耐热性的栽培种中有较大差异,在不耐热的不结球白菜品种(NHCC002)中未见该基因组成性表达;与38℃高温胁迫相比,4℃低温胁迫下诱导的BcHSP70-1表达量更高。

Primers were designed based on homologous sequences of previously reported Arabidopsis thaliana and Brassica napus L..BcHSP70-1 was isolated from Brassica campestris ssp.chinensis.The physical properties of BcHSP70-1 gene,together with subcellular localization and transmembrane domain were analyzed with bioinformatics.The results showed that：BcHSP70-1 contains 1 950 bp （ORF） nucleotides and encodes 649 amino acids,which is hydrophilic protein and localized in the cytoplasm.BcHSP70-1 has one intron and two exons,the intron is 324 bp.Comparison of orthologous exons and introns of HSP70-1 from different species showed that introns have more significant differences.Using real-time quantitative PCR analysis showed that BcHSP70-1 expressed differently among the different heat resistant cultivars.The heat sensitive cultivar （NHCC002） had no constitutive expression of the gene.The qRT-PCR results showed that,compared to 38 ℃ high temperature stress,BcHSP70-1 had higher expressed levels at 4 ℃ low temperature stress in non-heading Chinese cabbage.