Differential staining of Chironomus thummi giant chromosomes by treatment with acridine orange after mild acid hydrolysis

The fluorescence of Chironomus thummi giant chromosomes stained by acridine orange after mild acid hydrolysis (1N HC1 at 37°) has been studied. After very short hydrolysis as well as in control preparations (untreated by HCl) all chromosome regions show green fluorescence. After long hydrolysis (10 min) all chromosome bands fluoresce red. Intermediate time of hydrolysis (2 min) give in all transcriptionally inactive bands including centromeric ones red fluorescence whereas transcriptionally active puffing regions are green. The possible mechanisms of differential staining of individual chromosome regions are discussed. It is suggested that transcriptionally active chromosome regions are less susceptible to the action of acid or that the difference in chromatin stainability after acid-AO treatment is due to the difference in chromatin packing. The applicability of this method to the study of functional states of chromatin, at least in giant chromosomes, is discussed.