香蕉miR408b靶基因Chemocyanin的克隆及其在低温胁迫下的表达分析Cloning of miR408b target gene Chemocyanin and analysis of its expression under cold stress in banana
刘彦英;倪珊珊;项蕾蕾;孙雪丽;陈裕坤;赖钟雄;
摘要(Abstract):
【目的】研究香蕉中miR408b及其靶基因Chemocyanin在低温胁迫下的表达模式和调控机制。【方法】以三明野生蕉和‘天宝蕉’叶片为材料,利用RT-PCR和RACE技术克隆获得三明野生蕉中的MiChemocyanin基因(MiCHY)和‘天宝蕉’中的MaChemocyanin基因(MaCHY),利用在线网站对MiCHY进行生物信息学分析。【结果】MiCHY基因的cDNA全长为587 bp,开放阅读框381 bp,编码126个氨基酸蛋白,具有Plantacyanin结构域,属于超铜氧化还原蛋白家族。在ORF区的4~24 bp处有一处高度匹配miR408b的结合区(GCTCAGGGAAGGGGCAGTGCG)。RLM-5’RACE分析表明,miR408b主要在靶序列的第7~8个碱基之间剪切靶基因MiCHY的mRNA。序列比对发现,MaCHY基因比MiCHY基因多了82 bp的序列,符合内含子的GT-AG剪切,推测发生了内含子驻留的可变剪接。保守结构域分析表明MaCHY没有Plantacyanin结构域,但也属于铜氧化还原蛋白家族。qRT-PCR结果表明,低温胁迫下,冷敏感的‘天宝蕉’miR408b的表达量上升,MaCHY的表达量降低;在抗寒的三明野生蕉中,miR408b的表达量下降,MiCHY表达量上升,miR408b与MiCHY表达量呈负相关。【结论】miR408b及其靶基因MiCHY可能在香蕉响应低温胁迫中发挥重要作用。
关键词(KeyWords): 香蕉;miR408b;Chemocyanin基因;低温胁迫;表达分析
基金项目(Foundation): 国家重点研发计划(2019YFD1000901);; 国家现代农业产业技术体系(香蕉)专项资金(GARS-31-15);; 福建省高原学科建设经费(102/71201801101);; 福建农林大学科技创新专项基金(CXZX2018076)
作者(Author): 刘彦英;倪珊珊;项蕾蕾;孙雪丽;陈裕坤;赖钟雄;
Email:
DOI: 10.13925/j.cnki.gsxb.20190286
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