Mapping New Simple Sequence Repeat Markers in Hazelnut

New simple sequence repeat (SSR) markers (344) were developed in hazelnut (Corylus avellana L.) by mining genomic sequences of cv. ‘Jefferson’.  Fragments from 'Jefferson' that contained SSRs with motifs of 3-6 bases were initially aligned and compared in silico with Illumina genomic sequences from seven additional cultivars.  The alignments were visualized in Tablet software and inspected for variation in repeat number among the reads.  Of the 489 identified as polymorphic in Tablet, PCR amplification followed by separation on agarose gels confirmed polymorphism for 366 of them.  Twenty two sequences were identified as duplicates and removed.  Polymorphism in the remaining 344 was further validated by capillary electrophoresis after post-PCR multiplexing.  These new SSRs were added to the linkage map for hazelnut.  All 138 seedlings in a mapping population from a cross of OSU 252.146 x OSU 414.062 were amplified and scored for the 210 segregating SSR loci.  Of these, 130 segregated in a 1:1 ratio, 53 in a 1:1:1:1 ratio, and 57 in a 1:2:1 ratio.  The markers were assigned to linkage groups and maps were constructed using JoinMap 4.1.  All new SSRs were assigned to linkage groups, and 90% were placed on the map.  A dense linkage map with SSR markers will facilitate the study of traits, assignment of loci to linkage groups, mapping of additional markers, and marker-assisted selection.  Highly polymorphic and easy to score SSR markers that cover each LG at regular intervals will be useful for fingerprinting accessions and studying genetic diversity.