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. 2021 May 19;14(4):1642–1656. doi: 10.1111/1751-7915.13830

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© 2021 The Authors. Microbial Biotechnology published by John Wiley & Sons Ltd and Society for Applied Microbiology.

This is an open access article under the terms of the http://creativecommons.org/licenses/by-nc/4.0/ License, which permits use, distribution and reproduction in any medium, provided the original work is properly cited and is not used for commercial purposes.

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Fig. 6 — Exonuclease activity of Phi29 Pol and Phi29EL Pol.

A. Sequences and modifications of Oligos A, B and C. The lowercase ‘s’ denotes phosphorothioate.

B. Both polymerases (200 nM) were separately mixed with three oligos for 10 min. The reactions were analysed using a 20% TBE‐urea gel. 3′‐5′ proofreading exonuclease activity of both enzymes could degrade natural oligos (Oligo A) and 5′ phosphorothioate‐modified oligos (Oligo C) after a short time of incubation.

C. Specific activity assay. A 3′ to 5′ exonuclease activity assay was performed with Phi29 Pol or Phi29EL Pol according to the manufacturer’s instructions. Data are presented as means ± SD.