Loblolly pine karyotype using FISH and DAPI positive banding

The pines (Pinus, 2n = 2x = 24) include many commercially important timber species. Pinus spp. have 12 pairs of chromosomes, with 10 or 11 pairs of long metacentric chromosomes and one pair of short sub-metacentric chromosomes (Sax and Sax 1933). The pines have been studied extensively with conventional cytological techniques (Mergen 1958; Borzan and Papes 1978; MacPherson and Filion 1981; Schweizer 1980; Hizume et al. 1990). Molecular cytology, in situ hybridization (ISH), coupled with conventional cytological techniques can provide more accurate information about genomes (Heslop-Harrison 1991; Leitch and Heslop-Harrison 1992; Leitch et al. 1992). Well-spread metaphase chromosomes that are free of cell walls and cytoplasmic debris are a prerequisite for ISH. Since the chromosomes of pine are extremely large, wellspread metaphases are very difficult to obtain (Doudrick et al. 1995; Jacobs et al. 2000; Schmidt et al. 2000). We report a modified somatic chromosome preparation technique that was used to improve chromosome spreading and morphology in loblolly (Pinus taeda L.) and slash (P. elliottii Englm.) pines. Fluorescent ISH (FISH) was then used to locate 18S-28S ribosomal, 5S ribosomal, and telomeric DNA sites in these plant species, and to facilitate the development of karyotypes for each.