Acta Horticulturae Sinica ›› 2021, Vol. 48 ›› Issue (2): 377-388.doi: 10.16420/j.issn.0513-353x.2020-0306

• Research Notes • Previous Articles     Next Articles

Selection and Validation of Reference Genes for qRT-PCR Analysis of the Correlated Genes in Flower Pigments Biosynthesis Pathway of Anemone obtusiloba

MA Lulin1,2,*, DUAN Qing1,*, CUI Guangfen1, DU Wenwen1, JIA Wenjie1, WANG Xiangning1, WANG Jihua1,**(), CHEN Fadi2,**()   

  1. 1Flower Research Institute,Yunnan Academy of Agricultural Sciences,Yunnan Flower Breeding Key Lab,National Engineering Research Center for Ornamental Horticulture,Kunming 650205,China
    2College Horticulture of Nanjing Agricultural University,Nanjing 210095,China
  • Received:2020-09-27 Revised:2020-12-10 Online:2021-02-25 Published:2021-03-09
  • Contact: WANG Jihua,CHEN Fadi;


In order to screen the appropriate reference genes for qRT-PCR analysis of the flavonoids/anthocyanins biosynthetic pathway related genes of Anemone obtusiloba,eight traditional reference genes including polyubiquitin(UBQ),β-tubulin(β-TUB),aquaporin(AQP),actin(ACT),glyceraldehyde-3-phosphate dehydrogenase(GAPDH),histone(HIS),elongation factor 1-βEF-1β)and 60S ribosomal protein L13-1(RPL13)were selected as candidate reference genes based on the RNA-seq data of A. obtusiloba blue/white flowers. The expression of eight candidate reference genes were assessed by qRT-PCR in different tissues such as leaves,stems and blue/white different color flowers of A. obtusiloba,and the stability of them were analyzed by geNorm,NormFinder and BestKeeper programs. The relative expression of some pigments synthesis related genes in the biosynthetic pathway of flavonoids/anthocyanins were assessed to confirm the utility of the most stable reference gene. The results showed that UBQ was the most stable reference gene,while the stability of β-TUB was the lowest among all candidate reference genes. The qRT-PCR results of several pigments synthesis related genes using UBQ as the reference gene were in accordance with the results of RNA-seq. Thus,it was concluded that UBQ was the most suitable reference gene for gene expression analysis of flower pigments biosynthetic pathway in A. obtusiloba.

Key words: Anemone obtusiloba, qRT-PCR, reference gene, selection, stability, flavonoids/anthocyanins biosynthetic pathway

CLC Number: