Microsatellite isolation and characterization in pomegranate (Punica granatum L.)

Development of microsatellite markers has been an increasing trend in crop genetic studies because of their applicability in breeding programs. Here we report the development of SSRs in pomegranate using an enrichment method that makes use of magnetic beads. Enriched genomic libraries with AG and ATG microsatellite motifs were constructed, and 60 positive clones were detected by a colony PCR technique. Out of these, 32 (74.4%) were found to contain microsatellite sequences and 25 primer pairs were designed, of which 44% revealed polymorphisms, 48% showed monomorphic patterns and 8% generated weak amplification on a set of 20 pomegranate genotypes. Eleven microsatellite primers were selected to assess polymorphism in the set of genotypes. There were 44 alleles amplified over 13 loci, with an average of 3.38 alleles per locus. The mean polymorphism information content (PIC) value was 0.433 over 13 loci, which shows that the majority of the microsatellite loci are highly informative. Cluster analysis was able to separate genotypes based on their geographical distribution and type (i.e., wild or domestic). This study shows the isolation efficiency of the magnetic beads procedure, the abundance of microsatellites in pomegranate, and their potential usefulness in pomegranate genome mapping and for evaluating pomegranate genotypes.