Localization of Chromosomal Dna Sequences Homologous to Ribosomal Gene Type I Insertion Dna in Drosophila Melanogaster.

Abstract

This study was conducted to determine if DNA sequences homologous to type I insertion DNA present in ribosomal genes of D. melanogaster occur outside these genes. The approach used was: (1) to hybridize specific probe molecules in situ to salivary gl and polytene chromosomes and embryonic mitotic chromosomes, and (2) to analyze the resulting autoradiographic distribution of label over these chromosomes. Type I insertion DNA was isolated from recombinant plasmid pDmra56 and ('3)H-cRNA synthesized from this template was used as the experimental probe. Control in situ hybridization experiments included the use of: (1) ('3)H-28S rRNA recovered from the organism, (2) ('3)H-cRNA synthesized from "insertionless" rDNA derived from recombinant plasmid pDmrY22, and (3) ('3)H-cRNA synthesized from pMB9 plasmid DNA. Experimental results provide strong evidence that DNA sequences homologous to type I insertion DNA are not restricted to ribosomal genes. Autoradiographic patterns of labeling in both polytene and mitotic chromosomes corroborate these findings. For polytene chromosomes, the characteristic labeling pattern found was: (1) dense label over the nucleolus as expected, (2) dense label over the chromo-center including the (alpha) and (beta)-heterochromatic bases of each arm, and (3) light label over euchromatic regions of all arms (several b and s on the X and one on the 4th chromosome labeled consistently). For embryonic mitotic chromosomes, the characteristic labeling pattern was: (1) label over the NORs of both X and Y; (2) label over centromeric heterochromatin of sex chromosomes X and Y, 2nd and 3rd autosomes; (3) label over the telomeric regions of X, Y, 2 and 3; and (4) little or no label over euchromatin. It was also observed that: (1) one homologue of each pair of mitotic chromosomes labeled considerably higher than the other, and (2) the distribution of label in polytene chromosomes may vary developmentally. Label over interphasic nuclei was found to be consistent with results for the chromosomes. The label over cytoplasm and non-cytological areas provided a basis to which label over chromosomes could be compared. Although no functional role for the type I insertion DNA was found it is clear that with respect to chromosome organization, type I insertion DNA sequences occur outside ribosomal genes and appear to have a complex pattern of distribution.