A comparison of PCR-based markers for the molecular identification of Sphagnum species of the section Acutifolia

Jakub Sawicki, Monika Szczecińska


RAPDs, ISJs, ISSRs, ITS and katGs were applied to determine genetic relationships between common Sphagnum species of the section Acutifolia. Twenty populations were genotyped using ten ISJ primers, 12 pairs of katG primers, 10 ISSR and 10 RAPD primers, and a restriction analysis of ITS1 and ITS2. ISSR and katG markers revealed the greatest number of species-specific bands. An analysis of ITS1 and ITS2 regions with restriction enzymes also proved to be a highly effective tool for species identification.


Sphagnum; genetic similarity; molecular markers RAPD; ISJ; katG; ITS; ISSR

Full Text:



Goffinet B, Buck WR, Wall MA. Orthotrichum freyanum (Orthotrichaceae), a new epiphytic moss from Chile. Nova Hedw. 2007;131:1-11.

Plášek V, Sawicki J, Trávníčková V, Pasečná M. Orthotrichum moravicum (Orthotrichaceae), a new moss species from the Czech Republic. Bryologist. 2009;112(2):329-336. doi:10.1639/0007-2745-112.2.329.

Andrews AL, Britton EG, Emerson JT. Sphagnales – Bryales, Sphagnaceae/Andreaeaceae/Archidiaceae, Bruchiaceae, Ditrichaceae, Bryoxiphiaceae, Seligeriaceae. New York: New York Botanical Garden Pr Dept; 1913. (North American Flora, Series 1; vol 15).

Warnstorff C. Sphagnales-Sphagnaceae (Sphagnologia Universalis). Leipzig: Verlag von Wilhelm Engelmann; 1911. (Das Pflanzenreich; vol 42).

Hedenäs L. On the interdependence of some leaf characters within the Drepanocladus aduncus-polycarpus complex. J Bryol. 1996;19:311-324.

Cronberg N. Population structure and interspecific differentiation of the peat moss sister species Sphagnum rubellum and S. capillifolium (Sphagnaceae) in northern Europe. Plant Syst Evol. 1998;209(3-4):139-158. doi:10.1007/BF00985226.

Cronberg N, Natcheva R. Hybridization between the peat mosses, Sphagnum capillifolium and S. quinquefarium (Sphagnaceae, Bryophyta) as inferred by morphological characters and isozyme markers. Plant Syst Evol. 2002;234(1-4):53-70. doi:10.1007/s00606-002-0196-y.

Flatberg K, Thingsgaard K, Såstad S. Interploidal gene flow and introgression in bryophytes: Sphagnum girgensohnii × S. russowii, a case of spontaneous neotriploidy. J Bryol. 2006;28:27-37. doi:10.1179/174328206X90459.

Natcheva R, Cronberg N. Recombination and introgression of nuclear and chloroplast genomes between the peat mosses, Sphagnum capillifolium and Sphagnum quinquefarium. Mol Ecol. 2007;16(4):811-818. doi:10.1111/j.1365-294X.2006.03163.x.

Cronberg N. Isozyme evidence of relationships within Sphagnum sect. Acutifolia (Sphagnaceae, Bryophyta). Plant Syst Evol. 1996;203(1-2):41-64. doi:10.1007/BF00985236.

Krzakowa M, Melosik I, editors. The variability in Polish populations of Sphagnum taxa (Subsecunda section), according to morphological, anatomical and biochemical traits. Poznań: Bogucki Wydawnictwo Naukowe. 2000.

Cronberg N. Isozyme electrophoresis of Sphagnum: an analysis of methodology. Lindbergia. 1995;20(1):40-48.

Stenøien HK, Såstad SM. Genetic structure in three haploid peat mosses (Sphagnum). Heredity. 1999;82(4):391-400. doi:10.1038/sj.hdy.6884940.

Stenøien HK, Flatberg KI. Genetic variability in the rare Norwegian peat moss Sphagnum troendelagicum. Bryologist. 2000;103(4):794-801.

Gunnarsson U, Hassel K, Söderström L. Genetic structure of the endangered peat moss Sphagnum angermanicum in Sweden: a result of historic or contemporary processes? Bryologist. 2005;108(2):194-203. doi:10.1639/0007-2745(2005)108[0194:GSOTEP]2.0.CO;2.

Karlin EF, Boles SB, Shaw AJ. Resolving boundaries between species in Sphagnum section Subsecunda using microsatellite markers. Taxon. 2008;57:1189-2E.

Shaw AJ. Phylogeny of the Sphagnopsida based on chloroplast and nuclear DNA sequences. Bryologist. 2000;103(2):277-306.

Shaw AJ, Cox CJ, Boles SB. Phylogeny, species delimitation, and recombination in Sphagnum section Acutifolia. Syst Botany. 2005;30(1):16-33. doi:10.1600/0363644053661823.

Shaw AJ, Cox CJ. Variation in "biodiversity value" of peatmoss species in Sphagnum section Acutifolia (Sphagnaceae). Am J Bot. 2005;92(11):1774-1783. doi:10.3732/ajb.92.11.1774.

Szczecińska M, Sawicki J, Polok K, Hołdyński C, Zieliński R. Comparison of three Polygonatum species from Poland based on DNA markers. Ann Bot Fennici. 2006;43:379-388.

Bączkiewicz A, Sawicki J, Buczkowska K, Polok K, Zieliński R. Application of different DNA markers in studies on cryptic species of Aneura pinguis (Jungermanniopsida, Metzgeriales). Crypt Bryol. 2008;29(1):3-21.

Sawicki J, Zieliński R. Phylogenetic relationships between five Sphagnum species of the section Acutifolia based on DNA markers. Cas Slez Muz (A). 2008;57:259-264.

Polok K, Sawicki J, Kubiak K, Szczecińska M, Korzekwa K, Szandar K, et al. Evolutionary divergence within Pellia endiviifolia (Dicks.) Dum. from Poland. In: Prus-Głowacki W, Pawlaczyk E, editors. Variability and evolution – new perspectives. Poznań: Adam Mickiewicz University; 2005. p. 241-252.

Fiedorow P, Odrzykoski IJ, Szweykowska-Kulińska Z. Phylogenetic studies of liverworts using molecular biology techniques. Scientific Publications of the University of Silesia. 1998;1696:244-249.

Patterson E, Boles SB, Brook AJ. Nuclear ribosomal DNA variation in Leucobryum glaucum and L. albidum (Leucobryaceae): a preliminary investigation. Bryologist. 1998;101(2):272-277. doi:10.2307/3244204.

Sawicki J, Plášek V, Szczecińska M. Molecular evidence does not support the current division of Orthotrichum subgenus Gymnoporus. Plant Syst Evol. 2009;279(1-4):125-137. doi:10.1007/s00606-009-0153-0.

Vanderpoorten A, Goffinet B, Quandt D. Utility of the internal transcribed spacers of the 18S-5.8S-26S nuclear ribosomal DNA in land plant systematics with special emphasis on Bryophytes. In: Sharma AK, Sharma A, editors. Plant genome biodiversity and evolution: Lower groups. Enfield: Science Publishers; 2006. p. 385-407. (vol 2B).

Vekemans X, Jacquemart A. Perspectives on the use of molecular markers in plant population biology. Belg J Bot. 1997;129(2):91-100.

Pérez T, Albornoz J, Domínguez A. An evaluation of RAPD fragment reproducibility and nature. Mol Ecol. 1998;7(10):1347-1357.

Scott KM, Crandall-Stotler B. RAPD polymorphism as an indicator of population structure, breeding system, and speciation in Fossombronia. Bryologist. 2002;105(2):225-232.

Såstad SM, Flatberg KI, Cronberg N. Electrophoretic evidence supporting a theory of allopolyploid origin of the peatmoss Sphagnum jensenii. Nord J Bot. 1999;19(3):355-362. doi:10.1111/j.1756-1051.1999.tb01127.x.

Såstad SM, Stenøien HK, Flatberg KI, Bakken S. The narrow endemic Sphagnum troendelagicum is an allopolyploid derivative of the widespread S. balticum and S. tenellum. Syst Botany. 2001;26(1):66-74. doi:10.1043/0363-6445-26.1.66.

Zietkiewicz E, Rafalski A, Labuda D. Genome fingerprinting by simple sequence repeat (SSR)-anchored polymerase chain reaction amplification. Genomics. 1994;20(2):176-183. doi:10.1006/geno.1994.1151.

Kantety RV, Zeng X, Bennetzen JL, Zehr BE. Assessment of genetic diversity in dent and popcorn (Zea mays L.) inbred lines using inter-simple sequence repeat (ISSR) amplification. Mol Breeding. 1995;1(4):365-373. doi:10.1007/BF01248414.

Vanderpoorten A, Hedenäs L, Jacquemart A. Differentiation in DNA fingerprinting and morphology among species of the pleurocarpous moss genus, Rhytidiadelphus (Hylocomiaceae). Taxon. 2003;52(2):229-236. doi:10.2307/3647391.

Spagnuolo V, Terracciano S, Cobianchi RC, Giordano S. Taxonomy of the Hypnum cupressiforme complex in Italy based on ITS and trnL sequences and ISSR markers. J Bryol. 2008;30:283-289. doi:10.1179/174328208X300750.

Sawicki J, Plášek V, Szczecińska M. Molecular studies resolve Nyholmiella (Orthotrichaceae) as a separate genus. J Syst Evol. 2010;48(3):183-194. doi:10.1111/j.1759-6831.2010.00076.x.

Plášek V, Sawicki J. Is the hairy vaginula an diagnostic feature in the taxonomy of the genus Orthotrichum. Acta Soc Bot Pol. 2010;79(1):73-80.

Weining S, Langridge P. Identification and mapping of polymorphisms in cereals based on the polymerase chain reaction. Theoret Appl Genetics. 1991;82(2). doi:10.1007/BF00226215.

Szczecinska M, Sawicki J, Wasowicz K, Holdynski C. Genetic variation of the relict and endangered population of Chamaedaphne calyculata (Ericaceae) in Poland. Dendrobiology. 2009;62:23-33.

Song W, Henry RJ. Molecular analysis of the DNA polymorphism of wild barley (Hordeum spontaneum) germplasm using the polymerase chain reaction. Genet Resour Crop Evol. 1995;42(3):273-280. doi:10.1007/BF02431262.

Zieliński R, Polok K. Molecular evolution and plant taxonomy. In: Prus-Głowacki W, Pawlaczyk E, editors. Variability and evolution. Poznań: Adam Mickiewicz University; 2005. p. 37-56.

Krzakowa M, Pietkiewicz J, Polok K. Polimorfizm sekwencji genu katG u Calamagrostis arundinacea (L.) Roth. In: Jackowiak B, Celka Z, editors. Taksonomia, chorologia i ekologia roślin w dobie zagrożenia różnorodności biologicznej. Materiały konferencji naukowej dedykowanej Profesorowi dr hab. Waldemarowi Żukowskiemu z okazji 70-lecia urodzin. Poznań: Adam Mickiewicz University – Faculty of Biology – Department of Plant Taxonomy; 2005.

Polok K. Molecular evolution of the genus Lolium L. Olsztyn: Studio poligrafii komputerowej SQL; 2007.

Krzakowa M, Klama H, Wojtuń B. Interpopulation peroxidase variability in Sphagnum girgensohnii Russ. J Bryol. 1992;17(1):27-33.

Krzakowa M, Okonkwo N, Wojtuń B. Intraspecific diversity of Sphagnum russowii (Musci, Sphagnaceae) as revealed by peroxidase electrophoresis. Fragm Flor Geobot. 1995;40:447-451.

Pompanon F, Bonin A, Bellemain E, Taberlet P. Genotyping errors: causes, consequences and solutions. Nat Rev Genet. 2005;6(11):847-859. doi:10.1038/nrg1707.

Peakall R, Smouse PE. Genalex 6: genetic analysis in Excel. Population genetic software for teaching and research. Mol Ecol Notes. 2006;6(1):288-295. doi:10.1111/j.1471-8286.2005.01155.x.

Hey J, Wakeley J. A coalescent estimator of the population recombination rate. Genetics. 1997;145(3):833-846.

Zamocky M, Janecek S, Koller F. Common phylogeny of catalase-peroksydases and ascorbate peroxidases. Gene. 2000;256:169-182.

Krzakowa M, Melosik I, Hansjorg R. The taxonomic status of Sphagnum centrale (Musci, Sphagnaceae) as revealed by peroxidase diversity in relation to S. magellanicum and S. palustre. Fragm Flor Geobot. 1995;40:437-446.

Krzakowa M. The significance of cathodic peroxidases in the taxonomy of bryophytes. In: Welinder KG, Rasmussen SK, Panel C, Grepin H, editors. Plant peroxidases: biochemistry and physiology. Geneva: University of Copenhagen, University of Geneva; 1993. p. 213-219.

Boisselier-Dubayle MC, Jubier MF, Lejeune B, Bischler H. Genetic variability in the three subspecies of Marchantia polymorpha (Hepaticae): isozymes, RFLP and RAPD markers. Taxon. 1995;44(3):363-376. doi:10.2307/1223406.

Sawicki J, Nadolski R, Hościk M, Zieliński R. Identification of Conocephalum conicum and C. salebrosum (Hepaticophyta) based on DNA markers. Cas Slez Muz (A). 2008;57:63-80.

DOI: https://doi.org/10.5586/asbp.2011.017

Journal ISSN:
  • 2083-9480 (online)
  • 0001-6977 (print; ceased since 2016)
This is an Open Access journal, which distributes its content under the terms of the Creative Commons Attribution License, which permits redistribution, commercial and non-commercial, provided that the content is properly cited.
The journal is a member of the Committee on Publication Ethics (COPE) and aims to follow the COPE’s principles.
The journal publisher is a member of the Open Access Scholarly Publishers Association.
The journal content is indexed in Similarity Check, the Crossref initiative to prevent scholarly and professional plagiarism.
Polish Botanical Society