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. 2020 Dec 3;48(22):12858–12873. doi: 10.1093/nar/gkaa1163

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© The Author(s) 2020. Published by Oxford University Press on behalf of Nucleic Acids Research.

This is an Open Access article distributed under the terms of the Creative Commons Attribution License (http://creativecommons.org/licenses/by/4.0/), which permits unrestricted reuse, distribution, and reproduction in any medium, provided the original work is properly cited.

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Figure 2. — MTase activity requires single strands. Panels (A) and (C): M13 substrates stained with ethidium bromide. Panels (B) and (D): fluorograms of modification reactions using [H³]SAM. M13 SS: virion DNA substrate. M13 RF cut: DS replication intermediate RFI was digested following the labelling reaction for visual simplification; NdeI (Panels A and B) or NdeI+BamHI (Panels C and D). The substrates were treated with MTase proteins obtained with PURExpress in vitro transcription-translation (Panels A and B) or were partially-purified (Ni-NTA purification) proteins synthesized in vivo (Panels C and D). Lanes 1) empty pSAPv6 vector, 2) M.BceJIII WT (pAF9), 3) M.EcoGIX WT (pAF10) and 4) M.EcoGIX APPA variant (pAF11). H³ radiolabeled markers (M) are HindIII digested lambda DNA modified at A by M.EcoGII.