Quantitative determination of methylated CpG in satellite DNA I and in L1Rn DNA sequences extracted from rat kidney tissue and from rat kidney cell lines

The level of methylation of CpG has been determined in satellite DNA I and in an 1180-base-pair fragment of L1Rn DNA sequences extracted from rat kidney tissue and from two rat kidney cell lines, NRK B77 and NRK 52E. This determination was performed by HPLC analysis of 3'-deoxyribonucleoside monophosphates obtained after digestion of DNA labelled in vitro with alpha-32P]dGTP using DNA polymerase I. Results obtained show that L1 sequences are hypomethylated in rat cell lines (29.3% in NRK B77 and 18.6% in NRK 52E) when compared to the same fragment extracted from rat kidney tissue (47.6%). However, satellite DNA I in the cell lines is much less affected by the hypomethylation. Satellite DNA I purified from NRK B77 and NRK 52E contains 58.8% and 47.8% respectively of methylated CpG whereas it contains 62% of methylated CpG in rat kidney tissue. Therefore, the demethylation of CpG seems not to occur at random in these cell lines.