Format

Send to

Choose Destination
Int J Mol Sci. 2012;13(6):7586-93. doi: 10.3390/ijms13067586. Epub 2012 Jun 19.

Isolation and characterization of Simple Sequence Repeats (SSR) Markers from the moss genus Orthotrichum using a small throughput pyrosequencing machine.

Author information

1
Department of Botany and Nature Protection, University of Warmia and Mazury in Olsztyn, Plac Lodzki 1, Olsztyn 10-727, Poland; E-Mails: monika.szczecinska@uwm.edu.pl (M.S.); monika.milewicz@uwm.edu.pl (M.M.).

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.

KEYWORDS:

454 pyrosequencing; Orthotrichum; SSR markers

PMID:
22837714
PMCID:
PMC3397546
DOI:
10.3390/ijms13067586
[Indexed for MEDLINE]
Free PMC Article

Supplemental Content

Full text links

Icon for PubMed Central
Loading ...
Support Center