茶树低温胁迫下microRNA实时定量PCR内参基因的筛选

摘要

MicroRNA（miRNA）在基因的转录后调控上起到重要作用，对其表达水平的定量研究已成为一种常见实验，选择合适的内参基因是确保实时荧光定量PCR（qRT-PCR）准确性的先决条件。本研究以茶树为材料，挑选了9个候选内参基因，用 qRT-PCR技术检测它们在不同样本中的表达量，采用 BestKeeper和 geNorm软件进行表达稳定性综合分析。结果表明，一种茶树新 miRNA（PC-3p-222）在不同低温处理以及不同茶树组织中表达最为稳定，Ct平均值为22~23，丰度适中，可以作为茶树低温胁迫下miRNA qRT-PCR的内参基因，为miRNA定量研究工作提供参考。%MicroRNA (miRNA) plays an important role in post transcriptional regulation ofgene expression. The research of miRNA quantitative expression level has become more and more popular. Choosing a suitable reference gene is aprerequisiteof anaccurate real-time fluorescence quantitative PCR(qRT-PCR) assay.The expressionsof 9 candidate reference genes by qRT-PCR method in different tea (Camellia sinensis) sampleswere investigated and their stabilities wereanalyzed by BestKeeper and geNorm.The results showedthat a novel tea miRNA (PC-3p-222) was the most stable reference gene in different tea plant tissuesunder different cold temperature.The average Ct valuewas 22-23 which show edmoderate expression abundance.Therefore,itcan be used as the appropriate reference gene for miRNAqRT-PCR under cold stress,whichprovidesa good choice forthe quantitative of miRNA expression.