Analysis of calf-thymus satellite DNA: evidence for specific methylation of cytosine in C-G sequences

Abstract

Digestion of purified calf tymus satellite I (phi = 1.714 g/cm3) with a series of restriction enzymes shows that modification in this satellite occurs preferentially in the sequence C-G. This was also shown to be the case in the other satellites and in bulk chromosomal calf thymus DNA. Cloning of purified satellite I DNA in Escherichia coli makes sites, previously modified, available for cutting with certain restriction enzymes. All these 'new sites' contain the sequence C-G. High-resolution mass spectros-copy establishes that the satellites contain a low concentration of 5-methylcytosine. This infers that methylation which inhibits retriction enzyme cutting must occur preferentially in the sequence C-G. Hybridization of cRNA of cloned satellite I DNA with the satellites III (phi = 1.706 g/cm3) and IV (phi = 1.710 g/cm3) shows that there is no or little sequence homology between these satellites. Digestion of calf thymus satellite I DNA with endoR. EcoRI and subsequent hybridization studies with the fragments shows two EcoRI fragments in addition to the usual 1400-base-pair EcoRI repeat unit.