Validation of Reference Genes in a Population of Blueberry (Vaccinium corymbosum) Plants Regenerated in Colchicine.

ABSTRACT

For the first time we report the validation of reference genes in plants from a population of blueberry (Vaccinium corymbosum) clones cultured in vitro on a colchicine-supplemented medium. Nodal segment explants of the cultivar Duke were regenerated by organogenesis under different periods of colchicine 1 mg/L exposure (1, 2, 3, 5, 30 days). The clones selected for the study showed variability for phenotypic traits after 2 years of adaptation to field conditions, compared to plants of the donor genotype that were regenerated on a medium without colchicine. Vaccinium myrtillus (GAPDH) and Vaccinium macrocarpon (ATP1, NADH, RPOB and COX2) were used as reference genomes for primer design. The results show that colchicine treatments can cause genomic changes in blueberry plants. At the molecular level, exposure of plants to colchicine in early periods could promote an increase in gene expression of specific genes such as ATP1, COX2, GAPDH, MATK, NADH and RPOB. However, prolonged exposure (30 days) could decrease gene expression of the genes studied. For qPCR assays, the primers designed for ATP1, COX2, GAPDH and MATK genes showed high efficiency. In addition, the GAPDH, ATP1, NADH and COX2 genes showed high stability and could be recommended as potential reference genes for gene expression assays in Vaccinium.