大白菜快速碱化因子BrRALF1的克隆与特征分析

Cloning and Characterization of a Rapid Alkalinization Factor Gene BrRALF1 in Chinese Cabbage

利用cDNA-AFLP技术分析大白菜核雄性不育"两用系"AB02可育株(msms)和不育株(Msms)花蕾的基因表达谱,在可育株混合花蕾cDNA中扩增出1条特异表达条带TDF-3,通过RACE和RT-PCR技术克隆了该基因的cDNA全长序列。序列分析表明,该基因编码大白菜快速碱化因子,被命名为BrRALF1;BrRALF1全长cDNA序列为551bp,编码1个包含79个氨基酸残基的前体蛋白;前体蛋白N末端1～28位为信号肽,成熟蛋白含有保守的YIXY结构域、YXRGC结构域和4个半胱氨酸残基;预测BrRALF1蛋白含有5个翻译后修饰性位点。表达模式分析表明,BrRALF1在可育花的花丝、花药、雌蕊、萼片、花瓣中均表达,其中在花药中表达量最高;BrRALF1表达在不育株上受到强烈抑制。

Analysis of gene differential expression was performed by cDNA-AFLP in the genic male sterile line AB02 of Chinese cabbage.A differentially expressed cDNA fragment,TDF-3,was found only in fertile plants.The full-length cDNA of the gene related with TDF-3 was amplified by RACE and RT-PCR.The cDNA was 551 bp long,encoding rapid alkalinization factor of 79 amino acids with a signal tide of 28 amino acids.Mature BrRALF1 protein contained one conserved YIXY domain,one YXRGC domain and four cysteines which can form disulfide bonds.Five post-translational modification sites were predicted in mature BrRALF1 protein.BrRALF1 only expressed in big and middle floral buds of the fertile plants in AB02,not in male sterile plants.Tissue expression profile analysis indicated that BrRALF1 expressed in filaments,anthers,sepals,pistils,petals of fertile flowers,and the peak expression occurred in anthers.BrRALF1 expression was inhibited in male plants.