Methylation of linear dsDNA by pA-M.EcoGII inhibits multiple site-specific methylation-sensitive restriction endonucleases. The unmethylated DNA template was a 7-kb linear dsDNA, which was PCR amplified from pTXB1 plasmid. The DNA template was treated with commercial M.EcoGII, pA-M.EcoGII, and no enzyme. These treated DNA templates were each incubated with four restriction endonucleases (BamHI, EcoRV, PciI, PvuII). The BamHI is the m6A methylation insensitive enzyme, and the EcoRV, PciI, and PvuII are the m6A methylation-sensitive enzyme, with which the digestion could be blocked by corresponding m6A site. Our pA-M.EcoGII recombinant protein showed digestion inhibition on EcoRV, PciI, and PvuII digested samples, better than commercial M.EcoGII, as compared to untreated DNA template. DNA marker, 1 kb ladder, 250–10,000 bp.
