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Int. J. Mol. Sci. 2012, 13(6), 7586-7593; doi:10.3390/ijms13067586

Isolation and Characterization of Simple Sequence Repeats (SSR) Markers from the Moss Genus Orthotrichum Using a Small Throughput Pyrosequencing Machine

1
Department of Botany and Nature Protection, University of Warmia and Mazury in Olsztyn, Plac Lodzki 1, Olsztyn 10-727, Poland
2
Department of Genetics, University of Silesia, Jagiellonska 28, Katowice 40-032, Poland
3
Department of Biology and Ecology, University of Ostrava, Chittussiho 10, Ostrava 71000, Czech Republic
*
Author to whom correspondence should be addressed.
Received: 10 May 2012 / Revised: 12 June 2012 / Accepted: 13 June 2012 / Published: 19 June 2012
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Abstract

Here, we report the results of next-generation sequencing on the GS Junior system to identify a large number of microsatellites from the epiphytic moss Orthotrichum speciosum. Using a combination of a total (non-enrichment) genomic library and small-scale 454 pyrosequencing, we determined 5382 contigs whose length ranged from 103 to 5445 bp. In this dataset we identified 92 SSR (simple sequence repeats) motifs in 89 contigs. Forty-six of these had flanking regions suitable for primer design. We tested PCR amplification, reproducibility, and the level of polymorphism of 46 primer pairs for Orthotrichum speciosum using 40 individuals from two populations. As a result, the designed primers revealed 35 polymorphic loci with more than two alleles detected. This method is cost- and time-effective in comparison with traditional approaches involving cloning and sequencing. View Full-Text
Keywords: SSR markers; Orthotrichum; 454 pyrosequencing SSR markers; Orthotrichum; 454 pyrosequencing
This is an open access article distributed under the Creative Commons Attribution License (CC BY 3.0).

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MDPI and ACS Style

Sawicki, J.; Kwaśniewski, M.; Szczecińska, M.; Chwiałkowska, K.; Milewicz, M.; Plášek, V. Isolation and Characterization of Simple Sequence Repeats (SSR) Markers from the Moss Genus Orthotrichum Using a Small Throughput Pyrosequencing Machine. Int. J. Mol. Sci. 2012, 13, 7586-7593.

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