Plant Cell Rep (2003) 22:59–63 DOI 10.1007/s00299-003-0653-2 GENETICS AND GENOMICS F. Bogunic · E. Muratovic · S. C. Brown · S. Siljak-Yakovlev Genome size and base composition of five Pinus species from the Balkan region Received: 28 January 2003 / Revised: 2 May 2003 / Accepted: 3 May 2003 / Published online: 27 June 2003 # Springer-Verlag 2003 Abstract The 2C DNA content and base composition of five Pinus (2n=24) species and two Pinus subspecies from the Balkan region have been estimated by flow cytometry. P. heldreichii (five populations) and P. peuce (one population) were assessed for the first time, as also were subspecies of P. nigra (three populations—two of subspecies nigra and one of subspecies dalmatica) along with P. sylvestris, and P. mugo from the same region. The 2C DNA values of these Pinus ranged from 42.5 pg to 54.9 pg (41.7–53.810 9 bp), and the base composition was quite stable (about 39.5% GC). Significant differences were observed between two subspecies of P. nigra and even between two populations of subsp. nigra. The two other species (P. sylvestris and P. mugo) had 2C values of 42.5 pg and 42.8 pg, respectively, while that of P. peuce was 54.9 pg. These genome sizes are in accordance with published values except for P. sylvestris, which was 20% below estimates made by other authors. Keywords Pinus · Flow cytometry · Balkan area · Interspecific and intraspecific variations Introduction Genome size or DNA C-value is an important biodiversity character with many important practical implications (Bennett et al. 2000). Pinus species have often been investigated for their genome size (Miksche 1968; Dhir and Miksche 1974; Teoh and Rees 1976; Greilhuber 1988; Wakamiya et al. 1993; Valkonen et al. 1994; O’Brien et al. 1996; Murray 1998, Hall et al. 2000; Joyner et al. 2001), but there are still species of this genus whose DNA values are unknown. Different opinions exist as to the reality or not of constancy in genome size. In conifers, DNA amount has received particular attention since Miksche (1968) and Dhir and Miksche (1974) reported on DNA intraspecific variation. However, Teoh and Rees (1976), working on the same species, did not observed significant differences at the population level in Picea glauca, P. engelmani, and Pinus contorta. The usage of different methods, different internal standards, and dif- ferent plant tissues has given rise to various DNA content estimations for the same species. Greilhuber’s studies (1986, 1988) were a turning-point as they revealed the methodological source of many errors in genome size estimates. He established that “self-tanning” led to stoichiometric errors in the cyto-photometric determina- tion of nuclear DNA content and that these were caused by the type of fixation used prior to Feulgen staining in tannin-rich plants. In a detailed discussion of genome size in gymnosperms, Murray (1998) also ascribed reported differences to the methodological problems associated with the preparation of materials—especially Feulgen staining—and claimed that significant intraspecific vari- ation in DNA amount has not been found in gymnosperms despite serial measurements having been made with different methods in different laboratories. On the con- trary, Hall et al. (2000) detected variations in some tropical hard pines. Valkonen et al. (1994) also showed Communicated by M. Beckert F. Bogunic Faculty of Forestry, University of Sarajevo, Zagrebacka 20, 71000 Sarajevo, Bosnia and Herzegovina F. Bogunic · E. Muratovic · S. Siljak-Yakovlev Laboratory for research and protection of endemic resources, Department of Biology, Faculty of Sciences, University of Sarajevo, Zmaja od Bosne 35, 71000 Sarajevo, Bosnia and Herzegovina S. C. Brown Institut des Sciences du VØgØtal, CNRS UPR 2355, 91198 Gif-sur-Yvette, France S. Siljak-Yakovlev ( ) ) Ecologie, SystØmatique, Evolution, UniversitØ Paris-Sud, Bâtiment 360, UPRESA 8079, 91405 Orsay, France e-mail: sonia.yakovlev@ese.u-psud.fr Tel.: +33-1-69154698 Fax: +33-1-69154697