Evaluation of suitable reference genes for normalization of quantitative real-time PCR analysis in rice plants under Xanthomonas oryzae pv. oryzae--infection and melatonin supplementation

Abstract Exogenous melatonin (MT) was found to be an interesting tool for enhancing the resistance of rice to Xanthomonasoryzae pv. oryzae ( Xoo )-caused bacterial blight (BB). However, the accurate comparison of the expression levels across samples was a challenging task. In this work, the stability of 10 common used housekeeping genes under Xoo -infection and MT supplementation in rice was analyzed using quantitative real-time PCR (qRT-PCR), and algorithms geNorm, NormFinder and BestKeeper. Our results indicated that most reference genes remained stable in Xoo -infected rice plants, while a number of reference genes were affected by MT supplementation. Among all studied genes, the transcript levels of 18S ( 18S ribosomal RNA ) and UBC ( Ubiquitin-conjugating enzyme E2 ) remained unaltered by Xoo infection, while UBC and UBQ5 ( Ubiquitin 5 ) were the most stable genes when examining simultaneous Xoo -infection and MT supplementation, demonstrating that UBC is a suitable reference gene for qRT-PCR data normalization in rice under Xoo -infection and MT supplementation.