The isolation and characterisation of microsatellites in hop (Humulus lupulus L.)

Microsatellite markers have been increasingly used in crop genetic studies because of their high applicability in breeding programs and here we report on the development of such markers in hops. Six hop genomic libraries were constructed, enriched for microsatellite motifs GA, GT, ACA, AGA, CAG and ACTC. The enrichment level varied from 50% for GA repeats to 7.7% for tetranucleotide ACTC repeats, with an average enrichment of 32%. All classes of microsatellite repeat arrays were revealed in sequenced clones, the predominant class being interrupted arrays (51%), followed by perfect (42%) and compound (7%). Sixty microsatellite sequences were selected for primer design, of which 53% of primer pairs gave a simple banding pattern, 33% complex patterns and 13% of primer pairs generated no amplification on a set of four hop accessions. Twenty-five microsatellite markers were selected to assess polymorphism in 67 hop accessions. The co-dominant nature of microsatellite markers was demonstrated by pedigree and by segregation analysis. There were 265 alleles amplified over 25 loci, with an average of 10.6 alleles per locus. The highest average number of alleles was found within perfect repeats and at loci with the longest uninterrupted microsatellites arrays. The mean PIC value of 0.60 over 25 loci shows that the majority of the microsatellite loci are highly informative. A sequence similarity search resulted in the development of a plant universal marker for wide cross-species amplification. This study shows the isolation efficiency of the enrichment procedure, the abundance of microsatellites in hop, the high information content of the developed markers and their potential usefulness in hop genome mapping and individual identification.