扁桃AcCBF1转录因子的克隆及表达分析

Clone and expression analysis of AcCBF1 gene from almond

【目的】分离和克隆新疆栽培扁桃CBF1转录因子基因,了解其在扁桃响应环境胁迫和逆境相关激素诱导的表达情况。【方法】以新疆扁桃幼苗为试验材料,对其进行低温、干旱、盐及ABA处理,使用PCR技术从新疆扁桃‘双果’克隆得到CBF1转录因子基因,通过农杆菌将35S-Ac CBF1-GFP融合质粒转化洋葱表皮细胞后在激光共聚焦显微镜下观察结果,并通过实时荧光定量PCR技术分析了SG-Ac CBF1转录因子基因在不同非生物逆境胁迫下的表达情况。【结果】序列分析表明,该基因编码框为729 bp,编码242个氨基酸,蛋白质分子量为27.405 ku,命名为SG-Ac CBF1,GenBank登录号为KM245571。氨基酸序列同源比对分析证实,SG-Ac CBF1属于CBF转录因子家族基因。系统发育树分析表明,SG-Ac CBF1与甜樱桃Pa DREB1的亲缘关系最近。亚细胞定位分析显示,SG-Ac CBF1蛋白定位于细胞核中。实时荧光定量PCR分析结果表明,低温、干旱、盐及ABA均能诱导SG-Ac CBF1基因表达。【结论】SG-Ac CBF1转录因子具有核定位功能,并参与了植物对逆境胁迫的调控。由于该转录因子属于家族基因,因此其对环境胁迫响应的强弱还有待探讨。

【Objective】The aim of this study was to isolate of SG-AcCBF1 gene from Prunus communis and analyze its expression patterns in different tissues and with environmental stresses and stress-related hormones treatment.【Methods】The almond seedlings were used for low temperature,drought,high salt stress,and ABA treatment. SG-AcCBF1 was cloned from‘Shuangguo＇（P. communis） using PCR technology,recombinant plasmid 35S-AcCBF1-GFP was introduced into onion epidermal cells by Agrobacterium-mediated transformation,green fluorescence was monitored under a laser scanning confocal microscope. Finally,expression patterns after different treatment was studied by Real-time PCR.【Results】Sequence analysis revealed that the open reading frame of this gene was 729 bp in full-length and encodes a protein of 242 amino acids. Its relative molecular mass was approximately 27.405 ku,named SG- AcCBF1. Genbank Accession No. was KM245571. Phylogenetic analysis confirmed that SG-AcCBF1 belonged to CBF transcription factor gene family. The phylogenetic tree analysis showed that SG-AcCBF1 has closest genetic relationship with Prunus avium DREB1. Subcellular localization results showed that the SG-AcCBF1 was localized in the nucleus. The results of real-time PCR showed that low temperture,drought,salt,and ABA stresses induced the expression of SG-AcCBF1 dramatically.【Conclusion】Transcription factor SG-AcCBF1 had nuclear location function,and confirmed that SG-AcCBF1 was related tothe stress responsece. Since the SG-AcCBF1 belong to gene family,its function involed in environmentalstress response remains to be further research.