Efficiency of Inter-Simple Sequence Repeat PCR for Detecting Somaclonal Variation among Leaf-Culture-Regenerated Plants of Horseradish.

Abstract

Regenerated plants were obtained by culturing leaf explants of Akame and Aome strains of horseradish (Armoracia rusticana Gaertn.) on MS agar-solidified medium supplemented with 1 mg/l NAA + 1 mg/l TDZ. Extracted DNA from leaves of mother plants was amplified using random and simple sequence repeat (SSR) primers. The SSR primers generated a larger nwnber of scorable bands than the random ones, although inter-and intra-strain polymorphism in the banding pattern was detected by both SSR and random primers at alnrost the same level. An SSR primer, (CAGA)4, was selected for detecting somaclonal variation among regenerants from leaf-derived calli. Somaclonal variation among the regenerants was observed in the PCR products as well as in the leaf shape and color. Seventy one percent of the regenerants exhibited different banding patterns from those of their mother plant, whereas variation in the leaf shape and color was observed in 5 % of the regenerants. These findings suggest that inter-SSR PCR is an efficient tool for detecting somaclonal variation in horseradish. The frequency of the regenerants exhibiting different banding patterns from those of their mother plants and the number of variant bands per regenerant were higher in the Akame than in the Aome strain, suggesting that somaclonal variation in horseradish was affected by the genotype. Fifty percent and 60% of the missing and newly acquired bands, respectively, for the same fragment size were common to the regenerants originating from different leaf-derived calli, suggesting that there were mutable sites in the horseradish genome during the tissue culture process.