Conservation Genetics Resources

, Volume 5, Issue 2, pp 413–416 | Cite as

Isolation and characterisation of new microsatellite markers for the stonefly Brachyptera braueri comparing a traditional approach with high throughput 454 sequencing

  • Jutta GeismarEmail author
  • Carsten Nowak
Technical Note


We developed thirteen microsatellite markers for the stonefly Brachytpera braueri, a rare aquatic insect. We compared a “traditional” approach yielding an enriched library by cloning of positive host cells, hybridising and Sanger sequencing of target fragments with a 454 next generation sequencing approach. From a total of 881 fragments containing a repeat motif thirteen polymorphic loci were developed and tested for two populations. Number of alleles ranged from 4 to 20 and values of heterozygosity varied from 0.17 to 0.91 (HO) and 0.26 to 0.91 (HE). The markers are essential to investigate the colonisation potential of freshwater insects.


Short tandem repeats Parallel pyrosequencing Aquatic insects Dispersal 



We thank Ralph Küttner and Bodo Plesky for providing the samples. Funding comes from the Deutsche Forschungsgemeinschaft (HA3431/4-1). Additional funding is provided by the research funding program Landes-Offensive zur Entwicklung Wissenschaftlich-ökonomischer Exzellenz (LOEWE) of Hesse’s Ministry of Higher Education, Research, and the Arts.


  1. Brettfeld R, Bellstedt R (2000) Wiederfund der steinfliege brachyptera braueri (Klapálek, 1900) nach fast hundert Jahren in Thüringen (Insecta: Plecoptera). Abhandlungen und Berichte des Museums der Nat Gotha 21:111–112Google Scholar
  2. Holleley CE, Geerts PG (2009) Multiplex manager 1.0: a cross-platform computer program that plans and optimizes multiplex PCR. Biotechniques 46(7):511–517. doi: 10.2144/000113156 PubMedCrossRefGoogle Scholar
  3. Küttner R, Hohlmann M, Plesky B, Voigt H (2008) Zur verbreitung und ökologie von Brachyptera braueri (Klapálek, 1900) (Insecta: Plecoptera) in mitteldeutschland unter berücksichtigung weiterer plecoptera-arten des zeitigen frühjahres. Lauterbornia 63:31–50Google Scholar
  4. Larkin MA, Blackshields G, Brown NP, Chenna R, McGettigan PA, McWilliam H, Valentin F, Wallace IM, Wilm A, Lopez R, Thompson JD, Gibson TJ, Higgins DG (2007) Clustal W and clustal X version 2.0. Bioinformatics 23(21):2947–2948. doi: 10.1093/bioinformatics/btm404 PubMedCrossRefGoogle Scholar
  5. Margulies M, Egholm M, Altman WE, Attiya S, Bader JS, Bemben LA, Berka J, Braverman MS, Chen YJ, Chen ZT, Dewell SB, Du L, Fierro JM, Gomes XV, Godwin BC, He W, Helgesen S, Ho CH, Irzyk GP, Jando SC, Alenquer MLI, Jarvie TP, Jirage KB, Kim JB, Knight JR, Lanza JR, Leamon JH, Lefkowitz SM, Lei M, Li J, Lohman KL, Lu H, Makhijani VB, McDade KE, McKenna MP, Myers EW, Nickerson E, Nobile JR, Plant R, Puc BP, Ronan MT, Roth GT, Sarkis GJ, Simons JF, Simpson JW, Srinivasan M, Tartaro KR, Tomasz A, Vogt KA, Volkmer GA, Wang SH, Wang Y, Weiner MP, Yu PG, Begley RF, Rothberg JM (2005) Genome sequencing in microfabricated high-density picolitre reactors. Nature 437(7057):376–380. doi: 10.1038/nature03959 PubMedGoogle Scholar
  6. Meglecz E, Costedoat C, Dubut V, Gilles A, Malausa T, Pech N, Martin J-F (2010) QDD: a user-friendly program to select microsatellite markers and design primers from large sequencing projects. Bioinformatics 26(3):403–404. doi: 10.1093/bioinformatics/btp670 PubMedCrossRefGoogle Scholar
  7. Ronaghi M (2001) Pyrosequencing sheds light on DNA sequencing. Genome Res 11(1):3–11. doi: 10.1101/gr.11.1.3 PubMedCrossRefGoogle Scholar
  8. Rousset F (2008) GENEPOP’007: a complete re-implementation of the GENEPOP software for windows and linux. Mol Ecol Resour 8(1):103–106. doi: 10.1111/j.1471-8286.2007.01931.x PubMedCrossRefGoogle Scholar
  9. Rozen S, Skaletsky H (2000) Primer3 on the WWW for general users and for biologist programmers. Methods Mol Biol (Clifton, NJ) 132:365–386Google Scholar
  10. Van Oosterhout C, Hutchinson WF, Wills DPM, Shipley P (2004) MICRO-CHECKER: software for identifying and correcting genotyping errors in microsatellite data. Mol Ecol Notes 4(3):535–538. doi: 10.1111/j.1471-8286.2004.00684.x CrossRefGoogle Scholar
  11. Zuker M (2003) Mfold web server for nucleic acid folding and hybridization prediction. Nucleic Acids Res 31(13):3406–3415. doi: 10.1093/nar/gkg595 PubMedCrossRefGoogle Scholar
  12. Zwick P (1992) Stream habitat fragmentation—a threat to biodiversity. Biodivers Conserv 1(2):80–97. doi: 10.1007/bf00731036 CrossRefGoogle Scholar

Copyright information

© Springer Science+Business Media Dordrecht 2012

Authors and Affiliations

  1. 1.Department of River Ecology and Conservation, Conservation Genetics GroupSenckenberg Research Institutes and Natural History MuseumsGelnhausenGermany
  2. 2.Biodiversity and Climate Research Centre (BiK-F)Frankfurt am MainGermany

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