Genetic and epigenetic evaluations of citrus calluses recovered from slow-growth culture

The embryogenic callus of "Red Marsh" grapefruit was stored in vitro by slow growth culture method for one year, and survived with a significant weight increment over that period. The survivers regenerated somatic embryos more easily than the controls. Eight callus lines were used for genetic analyses. Although chromosome number variations were verified by cytological examination both in the controls and the stored samples, the ploidy level remained relatively stable during the storage period. Randomly amplified polymorphic DNA (RAPD) analysis was performed to detect DNA sequence variation. No difference in RAPD pattern was found with the 102 primers used. However, a methylation sensitive amplified polymorphism (MSAP) assay showed DNA methylation changes in the stored samples compared with the controls.