木薯MeASR3基因的克隆及表达分析

Cloning and Expression Analysis of MeASR3 Gene in Cassava

【目的】克隆木薯脱落酸、胁迫、成熟诱导蛋白(abscisic acid, stress, ripening-induced, ASR)的cDNA序列,检测该基因在ABA、H_2O_2、NaCl和甘露醇4种处理下的表达模式,为揭示ASR基因在木薯耐旱机制中的作用提供参考。【方法】以木薯KU50叶片总RNA反转录的cDNA为模板,利用PCR的方法扩增获得一个ASR家族基因,对其进行了氨基酸序列比对和进化树分析,并利用荧光定量PCR技术对该基因在多种处理下的表达模式进行分析。【结果】克隆得到的cDNA长度为459 bp,编码153个氨基酸,命名为MeASR3 (GeneBank登录号:XM_021744431.1)。序列比对结果表明MeASR3含有ASR家族蛋白的主要特征功能域,进化树显示MeASR3与橡胶树来源的ASR蛋白亲缘关系最近。荧光定量结果表明MeASR3在转录水平受到H_2O_2、甘露醇和高盐胁迫的诱导作用,同时受到ABA的抑制作用。【结论】克隆获得木薯MeASR3基因,该基因在转录水平响应ABA处理以及氧化、高盐和干旱胁迫,可能参与到ABA以及多种胁迫下的信号转导途径,可作为候选基因用于木薯耐旱机制的研究。

[Objective]To lay the foundation for further investigation on the specific functions and molecular regulation mechanisms of ASR, we isolated a ASR family gene from cassava, and investigated its expression patterns under various treatments.[Method] Using the cDNA re versed from the total RNA as the template, a ASR family gene was isolated from the cassava cultivar KU50 by RT-PCR method, and the com parison of amino acid sequences and phylogenetic analysis were carried out, as well as the expression patterns of ASR gene under various treatments were investigated by QPCR.[Result]The results showed that the cDNA of ASR gene was 459 bp in full length, and it encoded a putative protein with 153 amino acids, named MeASR3, its GeneBank accession number was XM_021744431.1. Multiple sequence align ment and phylogenetic analysis indicated that MeASR3 protein contained the conserved domains of ASR family and had a close genetic rela tionship with homologous proteins in Hevea brasiliensis. In addition, QPCR assay revealed that MeASR3 was induced by H2O2 , mannitol and salt stress, but inhibited by ABA at the transcriptional level.[Conclusion]MeASR3 responded to ABA, H2O2, mannitol and salt stress, which suggested that MeASR3 might play an important role in signal transduction pathways under ABA treatment and various abiotic stresses, and could be used as a candidate gene for further study of cassava drought tolerance mechanism.