5-Azacytidine-induced hypomethylation of tobacco HRS60 tandem DNA repeats in tissue culture

Abstract

The methylation status and 5-azacytidine-induced hypomethylation of CCGG sites within a family of tandemly organized, highly repeated DNA sequences of the Nicotiana tabacum L. nuclear genome (HRS60 family) were studied. As shown by in-situ hybridization experiments, the HRS60 family is clustered in a few regions of some tobacco chromosomes. The DNAs of leaf-derived calli, leaf-derived calli cultured on media with 5-azacytidine, and leaves were cleaved with restriction endonucleases differing in the sensitivity to the methylation of cytosine. After electrophoresis and Southern blotting they were hybridized with the HRS60 probe. We show that (i) CpG dinucleotides, and partially also CpCpG trinucleotides, of the HRS60 family are methylated in DNAs of the non-treated calli and leaves, and (ii) that these DNA repeats are sensitive to the action of a hypomethylating drug, 5-azacytidine.