Private workspaceQuick Protocol for Monarch® DNA Gel Extraction Kit (NEB #T1020) V.3

DOI
dx.doi.org/10.17504/protocols.io.bf7ujrnw
  • New England Biolabs1
  • 1New England Biolabs
Isabel Gautreau
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DOI: dx.doi.org/10.17504/protocols.io.bf7ujrnw
External link: https://www.neb.com/protocols/2015/12/08/quick-protocol-for-monarch-dna-gel-extraction-kit-t1020
Protocol Citation: New England Biolabs . Quick Protocol for Monarch® DNA Gel Extraction Kit (NEB #T1020). protocols.io https://dx.doi.org/10.17504/protocols.io.bf7ujrnwVersion created by Isabel Gautreau
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
Created: May 10, 2020
Last Modified: February 10, 2022
Protocol Integer ID: 36820
Keywords: Extracting, Monarch DNA Gel Extracting Kit, extracting dna, DNA, gel extraction
Abstract
This is the quick version of the Monarch® DNA Gel Extraction Kit Protocol (NEB #T1020). For the full protocol, please click here.
Guidelines
For detailed protocol and more information, visit www.neb.com/T1020

The full protocol is available here. 

A video protocol is available here.
Materials
MATERIALS
ReagentMonarch® DNA Gel Extraction KitNew England BiolabsCatalog #T1020
Safety warnings
Please refer to Safety Data Sheets (SDS) for health and environmental hazards.
Before start
  • Add 4 volumes of ethanol (≥ 95%) to one volume of DNA Wash Buffer. 
– For 50-prep kit, add 20 ml of ethanol to 5 ml of Monarch DNA Wash Buffer
– For 250-prep kit, add 100 ml of ethanol to 25 ml of Monarch DNA Wash Buffer
  • All centrifugation steps should be carried out at Centrifigation16.000 x g (~Centrifigation13.000 rpm ).
  • Please note: column holds 800 μl
Excise the DNA fragment from the agarose gel, taking care to trim excess agarose. Transfer to a 1.5 ml microfuge tube and weigh the gel slice. Minimize exposure to UV light.
Add 4 volumes of Gel Dissolving Buffer to the gel slice (e.g., 400 μl buffer per 100 μl or 100 mg agarose).
Incubate the sample between 37–55°C (typically Temperature50 °C ), vortexing periodically until the gel slice is completely dissolved (generally 5-10 minutes).
Note
The time that takes a gel slice to melt depends on the size of the slice, the temperature used in the incubation as well as the percent agarose used in the gel. The time recommended above should be used just as a guideline.

Note
For DNA fragments > 8 kb, an additional 1.5 volumes of water should be added after the slice is dissolved to mitigate the tighter binding of larger pieces of DNA (e.g., 100 μl gel slice: 400 μl Gel Dissolving Buffer: 150 μl water).

Insert column into collection tube and load sample onto the column. Spin for Centrifigation16000 x g, 00:01:00 , then discard flow-through.
Re-insert column into collection tube. Add Amount200 µL DNA Wash Buffer (with ethanol added)  and spin for Centrifigation16000 x g, 00:01:00 . Discarding flow-through is optional.
Repeat Step 5: Re-insert column into collection tube. Add Amount200 µL DNA Wash Buffer (with ethanol added)  and spin for Centrifigation16000 x g, 00:01:00 . Discarding flow-through is optional.
Transfer column to a clean 1.5 ml microfuge tube. Use care to ensure that the tip of the column does not come into contact with the flow-through. If in doubt, re-spin for Duration00:01:00 .
Add ≥ Amount6 µL DNA Elution Buffer to the center of the matrix. Wait for Duration00:01:00 , and spin for Centrifigation16000 x g, 00:01:00 to elute the DNA.
Note
Typical elution volumes are 6–20 μl. Nuclease-free water (pH 7–8.5) can also be used to elute the DNA. Yield may slightly increase if a larger volume of DNA Elution Buffer is used, but the DNA will be less concentrated. For larger size DNA (≥ 10 kb), heating the elution buffer to 50°C prior to use can improve yield.