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Fig. 1. Detection of negative supercoils of DNA on hsp70 in salivary gland. (A) Genomic organization of the hsp70 genes at 87A. Arrows denote the orientation of transcription. The hsp70 genes are flanked by specialized chromatin structures, scs and scs' (Udvardy et al., 1985). Restriction sites used for Southern analysis are indicated: X, XbaI; E, EcoRI; B, BglII. DNA fragments CR and DDS were used for hybridization probes. (B) Psoralen photocrosslinking to the hsp70 genes at 87A in salivary gland cells. Salivary glands from non-heat-shocked or heat-shocked larvae were incubated with indicated concentrations of psoralen. After photocrosslinking, genomic DNA was isolated, digested with XbaI, denatured, electrophoresed and analyzed by Southern hybridization using CR fragment as a probe. (C) Effect of previous nicking of DNA or inhibition of transcription on the psoralen photocrosslinking. Lanes 1-7: salivary glands from heat-shocked larvae were incubated with (lanes 2-7) or without (lane 1) psoralen and analyzed as in (B). Where indicated, the glands were irradiated with X-rays before (lane 3) or after (lane 5) photocrosslinking. The sample in lane 4 was mock treated without X-ray. The glands were treated with (lane 6) or without (lane 7) {alpha}-amanitin before incubation with psoralen. Lanes 8 and 9: salivary glands from non-heat-shocked (lane 8) or heat-shocked larvae (lane 9) were incubated with psoralen. Genomic DNA was digested with BglII and EcoRI, and analyzed as in (B) using DDS fragment as a probe.





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