火龙果miR164b-NAC调控关系在非生物胁迫下的应答作用  

hpo-miR164b involved in pitaya(Hylocereus polyrhizus)responses to abiotic stresses by regulating HpNAC expression

作　　者：余欢欢 杨鵾[1] 沈天娇 侯黔东 文晓鹏[1,2] YU Huan-huan;YANG Kun;SHENG Tian-jiao;HU Qian dong;WEN Xiao-peng(Key Laboratory of Plant Resource Conservation and Germplasm Innovation in Mountainous Region(Ministry of Education),Institute of Agro-bioengineering/College of Life Science,Guizhou University,Guiyang 550025,China;Institute for Forest Resources&Environment of Guizhou.College of Forestry,Guizhou University,Guiyang 550025,China)

机构地区：[1]贵州大学农业生物工程研究院/生命科学学院山地植物资源保护与种质创新教育部重点实验室,贵州贵阳550025 [2]贵州省森林资源与环境研究中心贵州大学林学院,贵州贵阳550025

出　　处：《生物技术》2021年第3期209-216,共8页Biotechnology

基　　金：国家自然科学基金项目(32060663);贵州省教育厅农业重大产业科学研究攻关项目(黔教合KY字[2019]021)。

摘　　要：[目的]探究火龙果中miR164b及其靶基因NAC在应答非生物胁迫下的表达模式和调控机制。[方法]利用PCR技术克隆hpo-miR164b前体序列及HpNAC序列。采用在线网站对hpo-miR164b、HpNAC进行生物信息学分析,运用qRT-PCR方法探究hpo-miR164b及HpNAC在高温42℃、低温4℃、100 mmol/L NaCl及0.38 mmol/L ABA等非生物胁迫下的转录变化规律。[结果]获得长度为334 bp的hpo-MIR164b茎环前体,该序列可形成典型的茎环结构,hpomiR164b和hpo-miR164b*位于两个相反臂上;HpNAC基因的开放阅读框为1092 bp,编码363个氨基酸,保守结构域NAM位于ORF的49-423 bp位置。qRT-PCR结果表明,在高温和高盐处理下,hpo-miR164b均上调表达;在低温处理下,hpo-miR164b先下调后上调;而在ABA处理下,hpo-miR164b上调表达。HpNAC的表达趋势则与hpo-miR164b相反。[结论]hpo-miR164对HpNAC基因具有负调控作用,且miR164-NAC在火龙果应答非生物逆境过程中发挥了重要作用。[Objective]To explore the expression pattern and regulatory mechanism of miR164b and its target gene NAC in dragon fruit under abiotic stress.[Method]The hairpin sequence of hpo-miR164b and the coding sequence(CDS)of HpNAC were cloned from pitaya.The online website was used for bioinformatics analysis of the cloned products.The qPCR quantification method was used to explore the changes of transcriptional levels of hpo-miR164b and HpNAC under different abiotic stresses such as high temperature,NaCl,and abscisic acid treatments etc.[Result]A 334 bp sequence of hpo-MIR164b was cloned from genomic DNA.Secondary structure analysis showed a characteristic and stable stem-loop structure,with hpo-miR164b and hpo-miR164b*located in 5’and 3’arms.ORF of HpNAC was 1092 bp in length,whose encoding 363 amino acids,and NAM was demonstrated to locate at 49-423 positions of ORF.The qPCR quantification analysis showed that hpo-miR164b was up-regulated as pitaya was subjected to the abiotic stresses such ashigh temperature and NaCl treatments etc.However,under ABA treatment,hpo-miR164b was up-regulated.Moreover,under low temperature,its expression was firstly repressed,followed by up-regulation.Conversely,the expression of HpNAC demonstrated the opposite trends as exposure to the above-tested stresses.[Conclusion]Taken together,hpo-miR164b plays a regulatory role in the control of HpNAC,which might influence the abiotic stress response of pitaya.

关 键 词：火龙果 miR164 HpNAC 非生物胁迫 表达分析 

分 类 号：Q756[生物学—分子生物学]