Molecular characterization of Cymbidium kanran cultivars based on extended random amplified polymorphic DNA (ERAPD) markers

Abstract

Fifty-four Cymbidium kanran cultivars from China, Japan and Korea were examined and analyzed by using the successive screening of 3'-end extended random primer amplified polymorphic DNA (ERAPD) markers to determine their molecular diversity and relationships. In ERAPD analyses, the strandspecific DNA sequence of direct oligonucleotide extension sequencing primers was independently read from each of the RAPD fragments without recourse to cloning or strand separation. Eight primers, identified from 80 original RAPD primers, produced strong repetitive polymorphic bands that were used in 3'-end-extended random primer amplified DNA marker analysis. The products of primers ACTGAACGCCCG + ACTGAACGCCGG and ACTGAACGCCC + ACTGAACGCC, linked to the same locus (2.5 - kb), were developed from the original ACTGAACGC RAPD primer; the products of this marker were more stable and specific than the original RAPD marker. Unweighted pair-group mean analysis (UPGMA) grouped them into two clusters based upon geographical traits. We demonstrated that the ERAPD technique is a powerful tool for cultivar identification and establishment of genetic relationships of cultivars in Cymbidium kanran.

Key words: Cymbidium kanran; genetic relationship; Extended random amplified polymorphic DNA (ERAPD)