望春玉兰实时定量PCR分析中内参基因的筛选

Selection of Internal Reference Genes in Real-time Quantitative PCR Analysis of Magnolia biondii

选择合适的内参基因是进行实时荧光定量PCR(qRT-PCR)分析的前提条件。本研究选取了两种花色望春玉兰(Magnolia biondii)在不同开花时期的花瓣作为研究材料,利用qRT-PCR测定了从望春玉兰花瓣转录组数据库中选取的10个候选内参基因(MbCYP,MbEF-1α,MbGAPDH,MbTEF,MbUBC,Mbα-TUB,Mbβ-TUB,MbUBQ,MbGBP,MbNADP)在花瓣中的表达量,并使用geNorm、NormFinder和BestKeeper软件对各候选内参基因表达量的稳定性进行评价。对3个分析软件的结果综合分析后得出,在两种花色望春玉兰不同开花时期的花瓣中,表达最稳定的候选内参基因依次为MbTEF、MbCYP和MbGAPDH,最不稳定的为MbUBC。研究表明,在后续研究中可选取MbTEF或是MbTEF和MbCYP二者组合作为内参基因。

Selection of suitable internal reference genes is a prerequisite for real-time quantitative fluorescence PCR(qRT-PCR)analysis.In this study,the petals of Magnolia biondii with 2 different colors at different flowering stages were selected as research materials.The expression levels of 10 candidate reference genes(MbCYP,MbEF-1α,MbGAPDH,MbTEF,MbUBC,Mbα-TUB,Mbβ-TUB,MbUBQ,MbGBP,MbNADP)in the petals of Magnolia biondii at different flowering stages were measured by qRT-PCR.The stability of the expression of candidate internal reference genes was evaluated by using geNorm,NormFinder and Best Keeper software.The comprehensive results of three analysis software showed that in the petals of Magnolia biondii with 2 different colors at different flowering stages,the most stable candidate reference genes were MbTEF,MbCYP and MbGAPDH,while the most unstable was MbUBC.The research indicated that MbTEF or the combination of MbTEF and MbCYP could be selected as reference gene in follow experiment.