Skip to main content
SpringerLink
Log in

C-value reassessment of plant standards: an image cytometry approach

  • Original Paper
  • Published:
Plant Cell Reports Aims and scope Submit manuscript

Abstract

Image cytometry (ICM) has been used to measure DNA 2C-values by evaluating the optical density of Feulgen-stained nuclei. This optical measurement is carried out using three basic tools: microscopy, digital video camera, and image analysis software. Because ICM has been applied to plants, some authors have remarked that studies should be performed before this technique can be accepted as an accurate method for determination of plant genome size. Based on this, the 2C-value of eight plants, which are widely used as standards in DNA quantifications, was reassessed in a cascade-like manner, from A. thaliana through R. sativus, S. lycopersicum, Glycine max, Z. mays, P. sativum, V. faba, to A. cepa. The mean 2C-values of all plants were statistically compared to the values reported by other authors using flow cytometry and/or ICM. These analyses demonstrated that ICM is an accurate and reliable method for 2C-value measurement, representing an attractive alternative to flow cytometry. Statistical comparison of the results also indicated Glycine max ‘Polanka’ as the most adequate primary standard. However, distinct authors have been advised that 2C DNA content of the reference standard should be close to that of the sample. As three further approaches also revisited the 2C-value of these eight plants, we have thus proposed a mean 2C-value for each eight species.

This is a preview of subscription content, log in via an institution to check access.

Access this article

Log in via an institution

Price excludes VAT (USA)
Tax calculation will be finalised during checkout.

Instant access to the full article PDF.

Institutional subscriptions

Fig. 1
Fig. 2

Similar content being viewed by others

References

  • Bennett MD, Leitch IJ (1995) Nuclear DNA amounts in angiosperms. Ann Bot 76:113–176. doi:10.1006/anbo.1995.1085

    Article  CAS  Google Scholar 

  • Bennett MD, Leitch IJ (1997) Nuclear DNA amounts in angiosperms-583 new estimates. Ann Bot 80:169–196. doi:10.1006/anbo.1997.0415

    Article  CAS  Google Scholar 

  • Bennett MD, Leitch IJ (2005) Plant genome size research: a field in focus. Ann Bot 95:1–6. doi:10.1093/aob/mci001

    Article  PubMed  CAS  Google Scholar 

  • Bennett MD, Leitch IJ (2011) Nuclear DNA amounts in angiosperms: targets, trends and tomorrow. Ann Bot 107:467–590. doi:10.1093/aob/mcq258

    Article  PubMed  CAS  Google Scholar 

  • Bennett MD, Smith JB (1976) Nuclear DNA amounts in angiosperms. Philos Trans R Soc B 274:228–274

    Article  Google Scholar 

  • Bennett MD, Smith JB, Heslop-Harrison JS (1982) Nuclear DNA amounts in angiosperms. Philos R Soc Lond B Biol 216:179–199

    Article  CAS  Google Scholar 

  • Bennett MD, Bhandol P, Leitch IJ (2000) Nuclear DNA amounts in angiosperms and their modern uses-807 new estimates. Ann Bot 86:859–909. doi:10.1006/anbo.2000.1253

    Article  CAS  Google Scholar 

  • Bennett MD, Leitch IJ, Price HJ, Johnston JS (2003) Comparison with Caenorhabditis (100 Mb) and Drosophila (175 Mb) using flow cytometry show genome size in Arabdopsis to be 157 Mb and thus 25% larger than the Arabdopsis Genome Initiative estimate of 125 Mb. Ann Bot 91:547–557. doi:10.1093/aob/mcg057

    Article  PubMed  CAS  Google Scholar 

  • Böching A, Nguyen VQH (2004) Diagnostic and prognostic use of DNA image cytometry in cervical squamous intraepithelial lesions and invasive carcinoma. Cancer Cytopathol 102:41–54

    Google Scholar 

  • Böcking A, Giroud F, Reith A (1995) Consensus report of ESACP task force on standardization of diagnostic DNA image cytometry. Anal Cell Pathol 8:67–74

    PubMed  Google Scholar 

  • Carvalho CR, Clarindo WR, Abreu IS (2011) Image cytometry: nuclear and chromosomal DNA quantification. In: Chiarini-Garcia H, Melo RCN (eds) Light microscopy, methods in molecular biology, vol 689. Humana Press, New York, pp 51–68

    Google Scholar 

  • Chieco P, Derenzini M (1999) The Feulgen reaction 75 years on. Hystochem Cell Biol 111:345–358

    Article  CAS  Google Scholar 

  • Chieco P, Junker A, van Noorden CJF (2001) Image cytometry. Microscopy handbooks, vol 46. Springer, New York

    Google Scholar 

  • Cruz CD (1997) Programa GENES—Aplicativo Computacional em Genética e Estatística. Editora UFV, Viçosa

    Google Scholar 

  • Doležel J, Bartoš J (2005) Plant DNA flow cytometry and estimation of nuclear genome size. Ann Bot 95:99–110. doi:10.1093/aob/mci005

    Article  PubMed  Google Scholar 

  • Doležel J, Greilhuber J (2010) Nuclear genome size: are we getting closer? Cytometry 77A:635–642. doi:10.1002/cyto.a.20915

    Article  Google Scholar 

  • Doležel J, Sgorbati S, Lucretti S (1992) Comparison of three DNA fluorochromes for flow cytometric estimation of nuclear DNA content in plants. Physiol Plantarum 85:625–631. doi:10.1111/j.1399-3054.1992.tb04764.x

    Article  Google Scholar 

  • Doležel J, Greilhuber J, Lucretti S, Meister A, Lysák MA, Nardi L, Obermayer R (1998) Plant genome size estimation by flow cytometry: inter-laboratory comparison. Ann Bot 82:17–26

    Article  Google Scholar 

  • Dunn JM, Bird-Lieberman EL, Coleman HG, Oukrif D, Arthur K, Lao-Sirieix P, Murray L, McManus D, Novelli MR, Fitzgerald R, Lovat LB (2011) Image cytometry DNA ploidy abnormalities are an independent risk factor for cancer progression in non-dysplastic barrett’s oesophagus and low grade dysplasia: results of the NIBR study. Gut 60:A172–A173. doi:10.1136/gut.2011.239301.367

    Google Scholar 

  • Greilhuber J (2005) Intraspecific variation in genome size in angiosperms: identifying its existence. Ann Bot 95:91–98. doi:10.1093/aob/mci004

    Article  PubMed  CAS  Google Scholar 

  • Greilhuber J (2008) Cytochemistry and C-values: the less-well-known world of nuclear DNA amounts. Ann Bot 101:791–804. doi:10.1093/aob/mcm250

    Article  PubMed  CAS  Google Scholar 

  • Greilhuber J, Ebert I (1994) Genome size variation in Pisum sativum. Genome 37:646–655

    Article  PubMed  CAS  Google Scholar 

  • Greilhuber J, Temsch EM (2001) Feulgen densitometry: some observations relevant to best practice in quantitative nuclear DNA. Acta Bot Croat 60:285–298

    Google Scholar 

  • Hardie DC, Gregory TR, Hebert PDN (2002) From pixels to picograms: a beginers’ guide to genome quantification by Feulgen image analysis densitometry. J Histochem Cytochem 50(6):735–749

    Article  PubMed  CAS  Google Scholar 

  • Haroske G, Baak JPA, Danielsen H, Giroud F, Gschwendtner A, Oberholzer M, Reith A, Spieler P, Böcking A (2001) Fourth updated ESACP consensus report on diagnostic DNA image cytometry. Anal Cell Pathol 23:89–95

    PubMed  CAS  Google Scholar 

  • Johnston JS, Bennett MD, Rayburn AL, Galbraith DW, Price HJ (1999) Reference standards for determination of DNA content of plant nuclei. Am J Bot 86:609–613

    Article  PubMed  CAS  Google Scholar 

  • Kindermann D, Hilgers C (1994) Glare correction in image cytometry. Anal Cell Pathol 6:165–180

    PubMed  CAS  Google Scholar 

  • Kron P, Suda J, Husband BC (2007) Applications of flow cytometry to evolutionary and population biology. Annu Rev Ecol Evol Syst 38:847–876

    Article  Google Scholar 

  • Marhold K, Kudoh H, Pak JH, Watanabe K, Spaniel S, Lihova J (2010) Cytotype diversity and genome size variation in eastern Asian polyploidy Cardamine (Brassicaceae) species. Ann Bot 105:249–264. doi:10.1093/aob/mcp282

    Article  PubMed  CAS  Google Scholar 

  • Mendonça MAC, Carvalho CR, Clarindo WR (2010) DNA content differences between male and female chicken (Gallus gallus domesticus) Nuclei and Z and W chromosomes resolved by image cytometry. J Histochem Cytochem 58:229–235. doi:10.1369/jhc.2009.954727

    Article  PubMed  Google Scholar 

  • Michaelson MJ, Price HJ, Ellison JR, Johnston JS (1991) Comparison of plant DNA contents determined by Feulgen microspectrophotometry and laser flow cytometry. Am J Bot 78:183–188

    Article  CAS  Google Scholar 

  • Moscone E, Baranyi M, Ebert I, Greilhuber J, Ehrendorfer F, Hunziker AT (2003) Analysis of nuclear DNA content in Capsicum (Solanaceae) by flow cytometry and feulgen densitometry. Ann Bot 92:21–29. doi:10.1093/aob/mcg105

    Article  PubMed  Google Scholar 

  • Motherby H, Pomjanski N, Kube M, Boros A, Heiden T, Tribukait B, Bocking A (2002) Diagnostic DNA-flow- vs. -image-cytometry in effusion cytology1. Anal Cell Pathol 24:5–15

    PubMed  CAS  Google Scholar 

  • Muñoz M, Riegel R, Seemann P (2006) Use of image cytometry for the early screening of induced autopolyploids. Plant Breed 125:414–416. doi:10.1111/j.1439-0523.2006.01230.x

    Article  Google Scholar 

  • Ochatt SJ (2008) Flow cytometry in plant breeding. Cytometry Part A 73A:581–598. doi:10.1002/cyto.a.20562

    Article  CAS  Google Scholar 

  • Praça MM, Carvalho CR, Novaes CRDB (2009) Nuclear DNA content of three Eucalyptus species estimated by flow and image cytometry. Aust J Bot 57:524–531. doi:org/10.1071/BT09114

    Article  Google Scholar 

  • Praça-Fontes MM, Carvalho CR, Clarindo WR, Cruz CD (2011) Revisiting the DNA C-values of the genome size-standards used in plant flow cytometry to choose the “best primary standards”. Plant Cell Rep 30:1183–1191. doi:10.1007/s00299-011-1026-x

    Article  PubMed  Google Scholar 

  • Puech M, Giroud F (1999) Standardisation of DNA quantitation by image analysis: quality control of instrumentation. Cytometry Part A 36A:11–17. doi:10.1002/(SICI)1097-0320(19990501)36:1<11:AID-CYTO2>3.0.CO;2-T

    Article  Google Scholar 

  • Schönswetter P, Suda J, Popp M, Weiss-Schneeweiss H, Brochmann C (2007) Circumpolar phylogeography of Juncus biglumis (Juncaceae) inferred from AFLP fingerprints, cpDNA sequences, nuclear DNA content and chromosome numbers. Mol Phylogenet Evol 42:92–103. doi:10.1016/j.ympev.2006.06.016

    Article  PubMed  Google Scholar 

  • Spooner DM, Van Den Berg RG, Rivera-Peña A, Velguth P, Del Rio A, Salas-Lópes A (2001) Taxonomy of Mexican and Central American Members of Solanum Series Conicibaccata (sect Petota). Syst Bot 26:743–756. doi:10.1043/0363-6445-26.4.743

    Google Scholar 

  • Suda J, Leitch JI (2010) The quest for suitable reference standards in genome size research. Cytometry Part A 77A:717–720. doi:10.1002/cyto.a.20907

    Article  CAS  Google Scholar 

  • Vilhar B, Dermastia M (2002) Standardization of instrumentation in plant DNA image cytometry. Acta Bot Croat 61:11–26

    CAS  Google Scholar 

  • Vilhar B, Greilhuber J, Koce JD, Temsch EM, Dermastia M (2001) Plant genome size measurement with DNA image cytometry. Ann Bot 87:719–728. doi:10.1006/anbo.2001.1394

    Article  CAS  Google Scholar 

  • Vogelbruch M, Rütten A, Böcking A, Kapp A, Kiehl P (2002) Differentiation between malignant and benign follicular adnexal tumours of the skin by DNA image cytometry. Br J Dermatol 146:238–243

    Article  PubMed  CAS  Google Scholar 

  • Voglmayr H (2000) Nuclear DNA amounts in mosses (Musci). Ann Bot 85:531–546. doi:10.1006/anbo.1999.1103

    Article  CAS  Google Scholar 

  • Xing S, Khanavkar B, Nakhosteen JA, Atay Z, Jöckel KH, Marek W, RIDTELC Lung Study Group (2005) Predictive value of image cytometry for diagnosis of lung cancer in heavy smokers. Eur Respir J 25:956–963. doi:10.1183/09031936.05.00118903

    Article  PubMed  CAS  Google Scholar 

  • Zonneveld BJM, Leitch IJ, Bennett MD (2005) First nuclear DNA amounts in more than 300 angiosperms. Ann Bot 96:229–244. doi:10.1093/aob/mci170

    Article  PubMed  CAS  Google Scholar 

Download references

Acknowledgments

We thank FAPEMIG (Fundação de Amparo à Pesquisa de Minas Gerais), CNPq (Conselho Nacional de Desenvolvimento Científico e Tecnológico) and FAPES (Fundação de Amparo à Pesquisa do Espírito Santo), Brazil, for their financial support. We would also like to thank the Dr. Jaroslav Doležel for generously providing the plant standards used in this study.

Author information

Authors and Affiliations

  1. Laboratório de Microscopia, Departamento de Produção Vegetal, Centro de Ciências Agrárias, Universidade Federal do Espírito Santo, CEP 29500-000, Alegre, ES, Brazil

    Milene Miranda Praça-Fontes & Wellington Ronildo Clarindo

  2. Laboratório de Citogenética e Citometria, Departamento de Biologia Geral, Centro de Ciências Biológicas, Universidade Federal de Viçosa, CEP 36570-000, Viçosa, MG, Brazil

    Carlos Roberto Carvalho

Authors
  1. Milene Miranda Praça-Fontes
    View author publications

    You can also search for this author in PubMed Google Scholar

  2. Carlos Roberto Carvalho
    View author publications

    You can also search for this author in PubMed Google Scholar

  3. Wellington Ronildo Clarindo
    View author publications

    You can also search for this author in PubMed Google Scholar

Corresponding author

Correspondence to Carlos Roberto Carvalho.

Additional information

Communicated by D. Zaitlin.

Rights and permissions

Reprints and permissions

About this article

Cite this article

Praça-Fontes, M.M., Carvalho, C.R. & Clarindo, W.R. C-value reassessment of plant standards: an image cytometry approach. Plant Cell Rep 30, 2303–2312 (2011). https://doi.org/10.1007/s00299-011-1135-6

Download citation

  • Received:

  • Revised:

  • Accepted:

  • Published:

  • Issue Date:

  • DOI: https://doi.org/10.1007/s00299-011-1135-6

Keywords

Search

Navigation