KARYOTYPE OF SLASH PINE (PINUS-ELLIOTTII VAR ELLIOTTII) USING PATTERNS OF FLUORESCENCE IN-SITU HYBRIDIZATION AND FLUOROCHROME BANDING

A karyotype and idiogram were prepared for slash pine (Pines elliottii Engelm, var, elliottii; 2n = 2x = 24) using 26 mitotic metaphase cells from root-tips of seedlings; each metaphase had 11 pairs of long metacentric chromosomes and one shorter pair of submetacentric chromosomes. Fluorescent in situ hybridization showed seven pairs of chromosomes with intercalary sites for genes of 18S-5.8S-25S rRNA (18S-25S rDNA) and one pair with a paracentromeric site. Probes of 5S rDNA localized a major site of hybridization on another pair of chromosomes, whereas two minor sites occurred on other chromosomes, one with and one without a site of 18S-25S rDNA. After in situ hybridization, chromosomes were stained with the GC-base-specific fluorochrome chromomycin A(3) (CMA). Positive bands at intercalary sites were detected only on two pairs of chromosomes, paracentromeric sites only on three pairs of chromosomes, and bands at both regions on four pairs, Many bands of CMA showed at sites of hybridization of 18S-25S rDNA but one major band occurred at a centromere without an rDNA site. After staining with 4',6-diamidino-2-phenylindole (DAPI), bands appeared at AT-rich intercalary and/or centromeric regions of nearly all chromosomes. A characteristic double band of DAPI occurred near the centromere of one pair of chromosomes. Patterns of fluorescence in situ hybridization and fluorochrome banding allowed us to identify all 12 pairs of chromosomes and to establish a standard karyotype. We propose that the numbers assigned to the chromosomes be used for homologous group designations in slash pine and as the framework for homoeologous group designations of chromosomes in other species of pine and perhaps other genera of conifers.