Characterization of extrachromosomal DNA in the flesh fly Sarcophaga bullata.

Abstract

The polytene pupal foot pad cells of the flesh fly Sarcophaga bullata contain numerous extrachromosomal DNA containing granules. We have determined both the origin and the nature of the DNA sequences present in these granules. Studies done with quinacrine staining of seven day old pupal foot-pad polytene nuclei showed that the granules fluoresced very brightly while the chromosomal bands to which the granules were attached did not. The only other highly fluorescent regions of the polytene karyotype were the centromeric heterochromatin of chromosomes C and E and several bands associated with the nucleolus of Chromsome A. When polytene nuclei were hybridized in situ with cRNA made from highly repetitive DNA, many of the granules positively labeled. Most of the label on these slides was concentrated on the centromeric heterochromatin of chromosomes C and E. Quinacrine staining of the foot-pad cells at very early stages of pupal development showed that when granules were present, they were always closely associated with the same two centromeric regions, those of chromosomes C and E. Since the highly repetitive DNA located in these centromeric regions is underreplicated, we conclude that the granules result from an extrusion process which takes place early during the polytenization of these cells. The chromosomal integrity of the centromeric heterochromatin of chromosomes C and E is apparently disrupted and repetitive sequences are dissociated from the chromosomes as DNA granules which then secondarily become associated with chromosomal bands throughout the nucleus.