Selection of Reference Genes for Expression Study in Pulp and Seeds of Theobroma grandiflorum (Willd. ex Spreng.) Schum

• Published in: PloS one 2016-08, Vol.11 (8), p.e0160646-e0160646
• Main Authors: Ferraz Dos Santos, Lucas, Santana Silva, Raner José, Oliveira Jordão do Amaral, Daniel, Barbosa de Paula, Márcia Fabiana, Ludke Falcão, Loeni, Legavre, Thierry, Alves, Rafael Moyses, Marcellino, Lucilia Helena, Micheli, Fabienne
• Format: Article
• Language: English
• Subjects: Analysis; Biochemistry; Biology and Life Sciences; Breeding; Cell division; Computer and Information Sciences; Confectionery; Cosmetics; Data interpretation; Dehydrogenases; DNA Primers; Economic importance; Flowers & plants; Fruits; Gene expression; Gene Expression Regulation, Plant; Genes; Genes, Plant; Genetic aspects; Genotypes; Glyceraldehyde-3-phosphate dehydrogenase; Molecular biology; Pennisetum glaucum; Plant breeding; Plant resistance; Plants (botany); Polymerase chain reaction; Product differentiation; Proteins; Pulp; Real-Time Polymerase Chain Reaction - methods; Real-Time Polymerase Chain Reaction - standards; Research and analysis methods; Seeds; Seeds - genetics; Stability analysis; Sterculiaceae - genetics; Studies; Theobroma grandiflorum
• ISSN: 1932-6203; 1932-6203
• DOI: 10.1371/journal.pone.0160646

Cupuassu (Theobroma grandiflorum [Willd. ex Spreng.] Schum) is a species of high economic importance in Brazil with great potential at international level due to the multiple uses of both its seeds and pulp in the industry of sweets and cosmetics. For this reason, the cupuassu breeding program focused on the selection of genotypes with high pulp and seed quality-selection associated with the understanding of the mechanisms involved in fruit formation. Gene expression is one of the most used approaches related to such understanding. In this sense, quantitative real-time PCR (qPCR) is a powerful tool, since it rapidly and reliably quantifies gene expression levels across different experimental conditions. The analysis by qPCR and the correct interpretation of data depend on signal normalization using reference genes, i.e. genes presenting a uniform pattern of expression in the analyzed samples. Here, we selected and analyzed the expression of five genes from cupuassu (ACP, ACT, GAPDH, MDH, TUB) to be used as candidates for reference genes on pulp and seed of young, maturing and mature cupuassu fruits. The evaluation of the gene expression stability was obtained using the NormFinder, geNorm and BestKeeper programs. In general, our results indicated that the GAPDH and MDH genes constituted the best combination as reference genes to analyze the expression of cupuassu samples. To our knowledge, this is the first report of reference gene definition in cupuassu, and these results will support subsequent analysis related to gene expression studies in cupuassu plants subjected to different biotic or abiotic conditions as well as serve as a tool for diversity analysis based on pulp and seed quality.