INCREASED INFORMATIVENESS OF RAPD ANALYSIS BY DETECTION OF MICROSATELLITE MOTIFS

The recently developed random-amplified microsatellite polymorphism (R AMPO) technique detects second-level amplification products that are u seful as molecular markers. In the first step of the procedure, genomi c DNA is amplified with a single arbitrary or microsatellite-complemen tary primer. PCR products are then electrophoretically separated, phot ographed, blotted and hybridized to a P-32-labeled microsatellite prob e. Autoradiography reveals highly reproducible, polymorphic, probe-dep endent fingerprints, which are different from the ethidium bromide sta ining patterns. In this paper, we report the successful application of various mono-, tri- and tetranucleotide repeat motifs as RAMPO probes . We also compare the efficiency of arbitrary vs. microsatellite prime rs for the generation of RAMPO patterns. Repeated rehybridization to d ifferent probes has expanded the information contained in a single ran dom-amplified polymorphic DNA (RAPD) gel at least fivefold. Pattern co mplexity varies with the length and sequence of the probe. Application of the technique to a genetic relatedness study in the genus Dioscore a (yam) yielded highly informative markers, mainly at an interspecific level.