A direct analysis of transcribed minichromosomes: All transcribed SV40 minichromosomes have a nuclease-hypersensitive region within a nuclease-free domain

• Published in: The EMBO journal Vol. 3; no. 12; pp. 2929 - 2936
• Main Authors: Choder, M, Bratosin, S, Aloni, Y
• Format: Journal Article
• Language: English
• Published: 01-01-1984
• ISSN: 0261-4189

A direct quantitative method was developed for detecting a small fraction of minichromosomes active in transcription and for analyzing their chromatin structure. For this, SV40 minichromosomes were incubated in vitro in the presence of ( alpha - super(32)P)UTP to allow elongation of the in vivo pre-initiated nascent RNA and thus labeling of the in vivo transcriptionally active molecules. In all molecules the cleavage occurred within the transcriptional regulatory region. In addition, analysis by electron microscopy revealed that SV40 minichromosomes having an nucleosome-free region (gap) are also more sensitive to the initial cleavage by both DNase I and Bg/I restriction enzyme than non-gapped molecules and that the initial DNase I cleavage occurs within the gap. Thus, at least in SV40, the nuclease-hypersensitive region is positively correlated with the existence of a gap.