May 16, 2020
Version 1
Protocol for removing ssDNA from dsDNA or RNA Samples (NEB #M0568) V.1
- 1New England Biolabs
- New England Biolabs (NEB)Tech. support phone: +1(800)632-7799 email: info@neb.com
External link: https://neb.com/protocols/2018/08/06/protocol-for-removing-ssdna-from-dsdna-or-rna-samples-m0568
Protocol Citation: New England Biolabs 2020. Protocol for removing ssDNA from dsDNA or RNA Samples (NEB #M0568). protocols.io https://dx.doi.org/10.17504/protocols.io.7ryhm7w
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: May 16, 2020
Protocol Integer ID: 28184
Abstract
The protocol described below will enable degradation of up to 20 pmol of a 25 nt ssDNA (~ 200 ng).
In order to degrade larger amounts of ssDNA or ssDNAs longer than 25 nt, we recommend adding more enzyme instead of extending the reaction time.
Users should note that ssDNAs longer than 25 nt may form secondary structures that hinder Thermolabile Exonuclease I activity.
Materials
MATERIALS
NEBuffer 3.1 - 5.0 mlNew England BiolabsCatalog #B7203S
Thermolabile Exonuclease INew England BiolabsCatalog #M0568
Nuclease-free WaterNew England BiolabsCatalog #B1500
Safety warnings
Please see SDS (Safety Data Sheet) for hazards and safety warnings.
Prepare a 20 µL reaction as follows:
| Sample containing ssDNA (up to 20 pmol of a 25-mer) | x μl | |
| NEBuffer 3.1 (NEB #B7203)* | 2 μl** | |
| Thermolabile Exonuclease I | 1 μl | |
| Nuclease-free water | to 20 μl*** |
*Most PCR buffers are compatible.
**No reaction buffer is necessary if enzyme is added to a PCR reaction.
***Scale larger reaction volumes proportionally.
Incubate at 37 °C for 00:04:00 .
Stop reaction by heat-inactivation at 80 °C for 00:01:00 .