Deoxyribonuclease II as a probe to sequence-specific chromatin organization: preferential cleavage in the 72 bp modulator sequence of SV40 minichromosome

Abstract

T prove sequence-specific chromatin structure of SV40 minichromosome, we further modified previously described hybridization mapping. Actually (i) the digestion patterns by two nucleases (micrococcal and DNAase II) were compared and (ii) the kinetics of nuclease digestion was analyzed from early time points when only a fraction of minichromosomes was cleaved once to longer digestions when oligo- and mononucleosomal bands appeared. DNAase II is shown to possess certain sequence specificity different from that of micrococcal nuclease. The major finding is that DNAase II preferentially cleaves the SV40 minichromosome at a distinct region of the genome known as 72 bp modulator element. Other hypersensitive sites are located near the replication origin and T-ag binding site II and also near BamHI site where termination of replication and "late" transcription occurs. Micrococcal nuclease splits the BglI-Hpaii region in a different manner.