Feb 16, 2022

Public workspaceLarge (Klenow) Fragment Blunting (M0210) V.2

DOI
dx.doi.org/10.17504/protocols.io.be6djha6
  • 1New England Biolabs
  • New England Biolabs (NEB)
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DOI: dx.doi.org/10.17504/protocols.io.be6djha6
External link: https://www.neb.com/protocols/2014/01/13/protocol-for-blunting-ends-by-3-overhang-removal-and-fill-in-of-3-recessed-5-overhang-ends-using
Protocol Citation: New England Biolabs 2022. Large (Klenow) Fragment Blunting (M0210). protocols.io https://dx.doi.org/10.17504/protocols.io.be6djha6Version created by New England Biolabs
License: This is an open access protocol distributed under the terms of the Creative Commons Attribution License,  which permits unrestricted use, distribution, and reproduction in any medium, provided the original author and source are credited
Protocol status: Working
We use this protocol and it’s working
Created: April 15, 2020
Last Modified: February 16, 2022
Protocol Integer ID: 35749
Keywords: Klenow Fragment, Large DNA Polymerase Fragment, Large Klenow Fragment, Large DNA Klenow Fragment
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)
Materials
MATERIALS
ReagentDNA Polymerase I Klenow Fragment - 200 unitsNew England BiolabsCatalog #M0210S
Safety warnings
Please refer to the Safety Data Sheets (SDS) for health and environmental hazards.
DNA should be dissolved in Concentration1 X NEBuffer or T4 DNA Ligase Reaction buffer supplemented with Concentration33 micromolar (µM) each dNTP.
Add 1 unit of Klenow per microgram DNA.
Pipetting
Incubate for Duration00:15:00 at Temperature25 °C .
Incubation
Stop reaction by adding EDTA to a final concentration of Concentration10 millimolar (mM) .
Pipetting
Heat for Duration00:20:00 at Temperature75 °C .
Incubation