油菜花粉发育相关基因RALFbn的克隆与表达分析

Cloning and Expression Analysis of RALFbn Related to Pollen Development in Brassica napus L.

根据美国NCBI数据库中快速碱化因子(RALF)类基因序列的已知信息,克隆了油菜的快速碱化因子基因RALFbn,对其核酸序列及预测蛋白进行了生物信息学分析,并在油菜多种组织内观测其表达情况。结果表明:(1)经克隆获得油菜RALFbn基因的cDNA序列全长为510bp,无内含子,编码79个氨基酸。(2)生物信息学分析发现,油菜RALFbn蛋白具有RALF类蛋白保守的"YIXY"区和4个保守的半胱氨酸残基,并且含有N豆蔻酰化位点、酪氨酸激酶Ⅱ磷酸化位点、蛋白激酶C磷酸化位点、和酪氨酸激酶磷酸化位点等多个生物活性位点,说明该蛋白在油菜中潜在的生理调节能力较为活跃。(3)RT-PCR检测RALFbn基因在油菜生殖器官中的表达结果发现,RALFbn主要在油菜雄蕊中表达,而在雌蕊、花瓣和萼片中没有表达。提示RALFbn基因极可能与油菜雄蕊中花粉的发育相关。

In this research,the rapeseed rapid alkalinization factor gene RALFbn was initially cloned from Brassica napus by the NCBI（National Center of Biotechnology Information） database information.Structure analysis of RALFbn,its predicted protein RALFbn and the expression of RALFbn were done.The results showed that：（1）The full-length cDNA of RALFbn in B.napus was 510 bp,no intron,encoded 79 amino acids.（2）The RALFbn protein had the conserved ＂YIXY＂ domain and four conserved Cysteine residues which were the characters of RALF family members.It also had N-myristoylation site,tyrosine kinase Ⅱ phosphorylation sit,protein kinase C phosphorylation site,tyrosine kinase phosphorylation site,which indicated an active physiological activity of the potential RALFbn.（3）RT-PCR analysis results showed that RALFbn mostly expressed in rape stamens,no expression in pistil,petals and sepals,indicating that RALFbn was closely related to pollen development of B.napus.