Jun 20, 2020
Version 1
Protocol for Exonuclease VIII, truncated (NEB #M0545) V.1
This protocol is a draft, published without a DOI.
- New England Biolabs1
- 1New England Biolabs
- New England Biolabs (NEB)Tech. support phone: +1(800)632-7799 email: info@neb.com
Protocol Citation: New England Biolabs 2020. Protocol for Exonuclease VIII, truncated (NEB #M0545). protocols.io https://protocols.io/view/protocol-for-exonuclease-viii-truncated-neb-m0545-7sbhnan
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: September 27, 2019
Last Modified: March 29, 2021
Protocol Integer ID: 28195
Keywords: exo VIII
Abstract
Exonuclease VIII, truncated efficiently degrades linear dsDNA from 5’ to 3’ direction, leaving supercoiled dsDNA intact.
* Note: For more precise results or partial digestions, we recommend titration of the enzyme to the intended substrate.
Materials
MATERIALS
EDTAThermo FisherCatalog #17892
Exonuclease VIII truncatedNew England BiolabsCatalog #M0545
Safety warnings
Please see SDS (Safety Data Sheet) for hazards and safety warnings.
Set-up the reaction as follows:
| COMPONENTS | 50 μl REACTION | |
| DNA | up to 1 µg | |
| NEBuffer 4 (10X) | 5 µl (1X) | |
| Exonuclease VIII (truncated) | 1 µl (10 units) | |
| Nuclease-free H2O (NEB #B1500) | up to 50 µl |
Incubate at 37 °C for 00:30:00 .
Stop reaction by adding EDTA to at least 11 millimolar (mM) .
Heat Inactivation 70 °C for 00:30:00 .
To clean up treated samples, we recommend using one of the following steps:
b. Running the reaction on an agarose gel, and then extracting the DNA (we recommend the Monarch Gel Extraction Kit, NEB #T1020), or
c. Performing a phenol/chloroform extraction followed by ethanol precipitation.