Automated screening and primer design of fish microsatellite DNA loci on pyrosequencing data

In recent years, massive sequencing approaches have allowed us to determine genomic structures of various organisms rapidly, raising novel applicability of the high-throughput sequence data obtained to various fields of biological studies. We present here a pipeline to search for microsatellite DNA and design PCR primers encompassing the microsatellites on genomic sequence data produced by 454 pyrosequencing. We tested this pipeline, called ‘Auto-primer’, on several fish genomic sequences and obtained many and various candidates for microsatellite DNA loci useful for detecting intraspecies genetic variability. This in silico search for microsatellite DNA is superior to conventional cloning methods, since any sequence patterns of repeat unit can be screened.