DETERMINATION OF RELIABLE REFERENCE GENES FOR REVERSE TRANSCRIPTION QUANTITATIVE REAL-TIME PCR FROM OIL PALM TRANSCRIPTOMES

Abstract

A set of reliable reference genes is essential for accurate quantification and interpretation of gene expression data using reverse transcription quantitative real-time polymerase chain reaction (RT-qPCR). In this study, Roche-454 RNA-seq reads from 27 libraries of various oil palm tissues were systematically analysed to identify a set of potential reference genes. Eleven candidate reference genes were identified from the transcriptome data. These genes, together with three oil palm reference genes previously identified for tissue culture samples (PD000380, PD00569, pOP-EA01332) and five classical housekeeping genes [glyceraldehyde-3-phosphate dehydrogenase (GAPDH), NAD5, TUBULIN, UBIQUITIN, ACTIN] were analysed across samples collected from various tissues from mature oil palm (leaf, root, endosperm, mesocarp, female flowers) and stages of tissue culture (non-embryogenic callus, embryogenic callus, polyembryoids, and shoots from polyembryoids). The expression levels of these genes were compared and evaluated using geNorm. Three genes (EgREF_5, EgREF_7 and EgREF_11) were found to be appropriate reference genes for normalising gene expression data from both mature plant and tissue culture samples.