蜡梅快速碱化因子基因CpRALF的克隆及表达分析  

Cloning and Expression Analysis of a Rapid Alkalinization Factor Gene CpRALF from Chimonanthus praecox

作　　者：王斌[1] 杨汶源[1] 孙晶晶[1] 马婧[1] 李名扬[1] 眭顺照[1] 

机构地区：[1]西南大学园艺园林学院,重庆市花卉工程技术研究中心,南方山地园艺学教育部重点实验室,重庆400716

出　　处：《园艺学报》2012年第12期2431-2438,共8页Acta Horticulturae Sinica

基　　金：国家自然科学基金项目(31070622);中央高校基本科研业务费专项资金项目(XDJK2013A004)

摘　　要：通过随机克隆测序的方法从蜡梅花cDNA文库中获得快速碱化因子基因,命名为CpRALF(GenBank登录号为:JQ217379)。扩增获得的DNA序列与cDNA序列一致,表明该基因不具有内含子,包含一个384bp的最大开放阅读框,编码一个长127个氨基酸的蛋白质。蛋白质序列同源比对发现其含有快速碱化因子特有的YIXY、YXRGC和4个保守的半胱氨酸以及双精氨酸结构。实时荧光定量PCR分析表明该基因在蜡梅茎、根和盛开的全花中表达量较高;在花发育过程中,蕾期和衰败期表达丰度高。在对蜡梅幼苗进行高温、低温、高盐、重金属4种非生物胁迫处理后,通过实时荧光定量PCR对CpRALF在幼叶中的表达量进行分析,结果表明高温、高盐和重金属能够促进该基因表达,而低温则抑制该基因的表达,且CpRALF对高盐与重金属胁迫的响应比对高温和低温胁迫的响应更为显著和迅速,表明该基因可能在蜡梅花发育以及非生物胁迫反应响应方面发挥作用。A new plant gene was identified by randomly cloning and sequencing from cDNA library constructed from Chimonanthus praecox flowers. The gene has an open reading frame （ORF） of 384 bp encoding a putative 127aa-polypeptide, and its mature form contains a conservative YIXY domain, a YXRGC domain and four cysteine residues characteristic to the RALF gene. This gene was thereafter named as CpRALF （GenBank accession No. JQ217379） . Real-time fluorescence quantitative PCR demonstrated that steady-state expression of CpRALF was higher in root and blooming flower compared to that in stem and leaf. And there is a lower expression level in cotyledon and tender leaves. During the lifespan, the expression of CpR,4LF was more abundant in bud stage and wither period than in other periods. The expression of CpRALF was inducible under various abiotic stresses including high temperature,salinity and heavy metals, while low temperature could inhibit the expression of CpRALE The transcriptional responses of CpRALF subject to salinity and heavy metal treatments were more sensitive and severe than other treatments. In conclusion, we propose that the CpRALF gene may play a significant role in Chimonanthus praecox flower development and abiotic stress responses.

关 键 词：蜡梅 快速碱化因子 基因克隆 表达分析 

分 类 号：S685.17[农业科学—观赏园艺]