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. 2022 Jun 1;5:68. doi: 10.1038/s42004-022-00685-5

Fig. 3. Evaluation of modified nucleotides 5 and 10 under PEX reaction conditions.

Fig. 3

Gel image (PAGE 20%) shows the results of PEX reactions with 3′-phos-dTTP 5 and 3’-phos-LNA-TTP 10 with 5′-FAM-labeled primer P1 (devoid of a 3′-phosphate moiety), template T1, and various DNA polymerases. Natural and modified triphosphates were at a final concentration of 200 µM. The following quantities of polymerases were used: Phusion (2 U), Hemo Klen Taq (8 reactions), Taq (5 U), Bst (8 U), Q5 (2 U), Therminator (2 U), Vent (exo−) (2 U): 60 °C, 1 h; Dpo4 (2 U), Deep Vent (2 U): 55 °C, 1 h; Kf (exo−) (5 U): 37 °C, 1 h. Negative control (T−): No polymerase added to the mixtures or reactions with only dATP and dCTP or dATP, dCTP, and dTTP only. Positive control (T+): with all natural nucleotides and Taq polymerase. All reactions were incubated at adequate reaction temperatures for 1 h. P represents unreacted, 5′-FAM-labeled primer.