Large (Klenow) Fragment Blunting (M0210)
New England Biolabs
Published: 2022-02-16 DOI: 10.17504/protocols.io.be6djha6
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Abstract
Protocol for blunting ends by 3' overhang removal and fill-in of 3' recessed (5' overhang) ends using DNA Polymerase I, Large (Klenow) Fragment (M0210)
Before start
Steps
1.
DNA should be dissolved in 1X or T4 DNA Ligase Reaction buffer supplemented with 33micromolar (µM) each dNTP .
2.
Add 1 unit of Klenow per microgram DNA.
3.
Incubate for 0h 15m 0s at 25°C.
4.
Stop reaction by adding EDTA to a final concentration of 10millimolar (mM).
5.
Heat for 0h 20m 0s at 75°C.
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