Characterization of the flanking regions of Zea mays microsatellites reveals a large number of useful sequence polymorphisms

Sequence characterization of the flanking regions of 52 sequence-tagged microsatellite loci and two gene fragments from 11 Zea mays inbred lines identified a total of 324 sequence polymorphisms. The sequence polymorphisms consisted of both single-nucleotide polymorphisms and insertions/deletions in a ratio of approximately two to one. The level of sequence variation within the flanking regions of microsatellites linked to expressed sequence tags was lower than microsatellites that were unlinked to expressed sequence tags. However, both types of microsatellites generated a similar number of sequence-based alleles across the 11 genotypes surveyed. In two out of 20 microsatellites examined in detail, evidence was found for size-based allele homoplasy. Conversion of the observed sequence polymorphisms into allele-specific oligonucleotides followed by covalent binding to glass slides allowed the sequence polymorphisms to be used in a simple hybridization-based genotyping procedure. This procedure enabled us to discriminate between different inbred lines and allowed variations within a single inbred to be identified. The sequence information presented in this report could be used as a starting point for other programmes in the further development of a non-gel based, multi-locus, multi-allele screen for large-scale maize genotyping.