Cloning and selection of reference genes for gene expression studies in Ananas comosus
AbstractFull length mRNA sequences of Ac-β-actin and Ac-gapdh, and partial mRNA sequences of Ac-18SrRNA and Ac-ubiquitin were cloned from pineapple in this study. The four genes were tested as housekeeping genes in three experimental sets. GeNorm and NormFinder analysis revealed that β-actin was the most appropriate reference gene for qPCR analysis of callus under induction conditions and in different tissue types, meanwhile, 18SrRNA was the most stable reference gene during organ development. Gapdh was the most unstable gene in all tested experimental sets. Transcript level analysis result of AcSERK1 in stressed callus normalized by β-actin and 18SrRNA further confirmed that reference genes selected in this study were suitable for transcript level analysis of pineapple. The expression pattern of AcSERK1 during somatic embryogenesis normalized by β-actin coincided with the cytological features of calluses during somatic embryogenesis. These results will enable more accurate and reliable normalization of qPCR results for transcription analysis in pineapple.
Keywords: Reference genes, qPCR, pineapple, geNorm, NormFinder
African Journal of Biotechnology Vol. 11(29), pp. 7424-7433, 10 April, 2012