甜瓜ADF7基因的克隆与生物信息学分析

Cloning and Bioinformatics Analysis of ADF7 Gene in Melon(Cucumis melo L.)

肌动蛋白解聚因子(actin-depolymerizing factors,ADFs)通过剪切和解聚微丝来调控微丝组织和动态,被认为是肌动蛋白变化的重要调控因子。通过RT-PCR方法扩增得到甜瓜ADF7(CmADF7)基因序列,进行生物信息学分析和组织表达分析。结果表明CmADF7基因的开放阅读框(ORF)为414 bp,编码137个氨基酸,蛋白质理论分子量15.82102 kD,等电点5.32。生物信息学预测该蛋白是不具有跨膜结构的疏水性蛋白,亚细胞定位于细胞质中。基因结构分析显示CmADF7包含3个外显子,2个内含子,其中第1位外显子仅包含起始密码子ATG。结构域分析表明CmADF7具有高度保守的ADF-H结构域,其氨基酸序列为12~137位。同源性与系统进化分析证明CmADF7与同属葫芦科的西瓜的同源ADF亲缘关系较近。利用qRT-PCR分析发现CmADF7基因主要在花中表达,其他组织中检测虽有表达,但表达量都很低。序列分析表明,CmADF7启动子上存在一些参与光响应、胁迫应答及激素调节等相关顺式作用元件。新的甜瓜ADF蛋白结构和组织表达研究,将为进一步探索CmADF7蛋白功能和发掘其他CmADFs奠定基础。

Actin-depolymerizing factors(ADFs)are considered as the main regulatorsin organizing dynamics of filamentous actin(F-actin)by severing and depolymerizing F-actin.Here,ADF7 sequence in melon was cloned through RT-PCR.Bioinformatical analysis showed that the open reading frame(ORF)of CmADF7 was 414 bp,and encoded 137 amino acids.It weighted 15.82102 kD with a predicted pI of 5.32.It might be a hydrophobic protein without transmembrane structure locatedin the cytoplasm.Gene structure analysis reveals that it contained 3 exons and 2 introns,and the first exon only contained the start codon ATG.CmADF7 had highly conserved ADF-H domain(D12 to F137).The phylogenetic tree showed that watermelon,the same cucurbit family to melon,possessed the closest phylogenetic relationship and the highest homology.qRT-PCR results showed that the CmADF7 gene was mainly expressed in flowers,and the expression was also detected in the other tissues,but the expression level was very low.Sequence analysis revealed several putative cis-acting regulatory elements associated with the light signaling,stress and hormone response in CmADF7 promoter on the Plant CARE database.Our results provide evidences of a new CmADFs,and contribute to further exploring the function of CmADF7 and other ADFs in melon.