The moss Physcomitrella patens is an emerging model in comparative plant science. At present, the Physcomitrella genome is sequenced at the Joint Genome Institute (USA). In this study we present our results on the development of expressed sequence tag-derived microsatellite markers for Physcomitrella patens, their classification and applicability as genetic markers on the intra- as well as on the interspecies level. We experienced severe restrictions to compare our results on Physcomitrella with earlier studies for other plant species due to varying microsatellite search criteria and a limited selection of analysed species. As a consequence, we performed a side by side analysis of expressed sequence tag-derived microsatellites among 24 plant species covering a broad phylogenetic range and present our results on the observed frequencies.
We identified 3,723 microsatellites using the software MISA in a non-redundant Physcomitrella expressed sequence tag database comprising more than 37 megabases of nucleotide information. For 2,951 microsatellites appendant primer sequences have been derived. PCR of 376 microsatellites yielded 88 % successful amplicons and over 30 % polymorphisms between two Physcomitrella accessions. The polymorphism information content of 64 microsatellites based on 21 different Physcomitrella accessions was comparably high with a mean of 0.47 +/- 0.17. Of the 64 Physcomitrella microsatellite markers, 34 % respectively 79.7 % revealed cross-species applicability in two closely related moss species.
In our survey of two green algae, two mosses, a fern, a fern palm, the ginkgo tree, two conifers, ten dicots and five monocots we detected an up to sevenfold variation in the overall frequency with a minimum of 37 up to maximal 258 microsatellites per megabase and a high variability among the different microsatellite class and motif frequencies. Numerous species-specific microsatellite frequencies became evident and several deviations to earlier reports were ascertained.
With the Physcomitrella microsatellite marker set a valuable tool has been made available for further genetic and genomic applications on the intra- as well as on the interspecies level. The comparative survey of expressed sequence tag-derived microsatellites among the plant kingdom is well suited for a classification of future studies on plant microsatellites.