Abstract
The in situ detection of specific DNA sequences by fluorescence in situ hybridization (FISH) is a widely used tool for physical mapping, which is important to understand genome structure and organization. For species with small chromosomes, detection of the position of marker sequences is useful to aid chromosome identification and classification. Primed in situ labeling (PRINS) comprises an alternative to FISH, but its specificity and sensitivity may be influenced by species-specific factors, as chromosome length and chromatin structure. To overcome these obstacles, we designed a reproducible PRINS protocol for physical mapping of 5S rDNA in Eucalyptus dunnii Maiden and Zea mays L. Slides containing mitotic metaphases were submitted to one cycle of PRINS using 5S rDNA primers. Clear signals for 5S rDNA were observed in metaphase chromosomes of both species. In E. dunnii, 5S rDNA was mapped on the pericentromeric region of the short arm of chromosome 5. For Z. mays, the signal was detected in the terminal region of the long arm of chromosome 2. Although chromosome structure may influence the PRINS resolution, a high signal/background ratio was obtained for both species. In Z. mays, clear 5S rDNA signals were obtained by PRINS and FISH. Nonetheless, the former was superior regarding the time required for signal detection. Therefore, PRINS has the potential to aid plant genome studies by allowing fast detection of repetitive sequences, even in species with small chromosomes. Moreover, this is the first report of specific sequence mapping in Eucalyptus by PRINS, contributing to physical mapping progress in this genus.
