Determination of Inheritance of Methylated DNA

Abstract

I have studied the maintenance of methyl groups at the HpaII sites of autonomously functioning DNA upon transformation into mammalian cells. DNA molecules, containing the SV40 origin, were methylated in vitro with the bacterial modification enzyme HpaII methylase, which methylates the internal cytosine of the 5’-CCGG-3’ sequence. The DNAs were then transfected into a SV40 monkey cell line (COS) and extracted 48 hours later to determine methylation. The question of whether the DNAs were replicated was also asked. Restriction enzymes and Southern blotting were used to determine both methylation and replication. It was found that the DNAs replicated by COS cells lost their methylation, while the DNAs not replicated maintained their methylation. As a result, it is suggested that COS cells not be used in experiments requiring a specific methylation pattern to be maintained in transfected DNA.