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Crystallization

NC Nicholas Chim
RM Roman A. Meza
AT Anh M. Trinh
KY Kefan Yang
JC John C. Chaput
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Bst, at a final concentration of 4 mg mL−1, was incubated with 3 molar equivalents of the P/T duplex at 37 °C for 30 min, followed by a second incubation with 10 M excess of dTTP and 10 mM manganese chloride at 37 °C for 30 min to obtain the starting structure (0 min, 6DSY). 24–well plate hanging drop trays were used to optimize crystal growth over a range of ammonium sulfate and MPD concentrations, with each drop containing 1 μL of sample mixed with 1 μL of mother liquor over 500 μL of mother liquor in every well. Trays were stored in the dark at room temperature and growth of Bst binary crystals was generally observed after 2 days.

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