Optimizing RAPD Markers for Ginseng Genomic DNA Analysis

Abstract

Random amplified polymorphic DNA (RAPD) may have utility as genetic markers facilitating selection in ginseng crop improvement. This experiment determined chemical buffer and root tissue-type combinations that yield repeatable bands. The results allow further experiments using RAPD markers for estimating the genetic distance between ginseng landraces, selection for crop improvement, and extensive fingerprinting for use in determining the origin of tissue samples. This experiment determined mean band yields for all combinations of dry, fresh, and powdered root with cetyltrimethylammonium bromide, potassium/sodium ethyl xanthogenate, and urea buffers. The buffers were applied in replication to the tissue-types with other extraction protocol factors constant. Replications were amplified four times with four different primers using constant PCR and agarose gel electrophoretic protocols. Distinct bands were counted in each replication, and the summation of the replication repeats considered an observation. Least squares means for several response variables were analyzed. The most significant difference found was between buffers. The buffers ctab and urea were productive, and the pex was not. Significant difference was found when buffers were crossed with tissue. The applications of urea to fresh root, ctab to dry root, urea to dry root, and ctab to powdered root were productive. Based on these results we conclude 1) urea and ctab are productive when applied to all tissue-types, 2) dry root, which is easily collected and stored, yields sufficient DNA for analysis, and 3) powdered root, often the form of commercial products that might be tested for genetic origin, will yield sufficient DNA for analysis.