May 29, 2024

Qubit4 Fluorometer Protocol: Invitrogen dsDNA HS Assay Kit (REF 32854)

DOI
https://dx.doi.org/10.17504/protocols.io.81wgbz6dngpk/v1
  • 1University of Georgia- Holly Bik;
  • 2University of Georgia;
  • 3Department of Marine Sciences, University of Georgia;
  • 4UGA
  • Bik Lab UGA
Jenna Brown
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DOI: https://dx.doi.org/10.17504/protocols.io.81wgbz6dngpk/v1
Protocol Citation: Jenna Brown, Mirayana Marcelino Barros, Tiago Pereira, Alejandro De Santiago Perez, Hunter Powell 2024. Qubit4 Fluorometer Protocol: Invitrogen dsDNA HS Assay Kit (REF 32854). protocols.io https://dx.doi.org/10.17504/protocols.io.81wgbz6dngpk/v1
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: May 02, 2024
Last Modified: May 29, 2024
Protocol  Integer ID: 99081
Keywords: qubit4 fluorometer protocol, invitrogen dsdna hs assay kit, qubit4 protocol, dsdna concentrations in sample, dsdna concentration, protocol
Funders Acknowledgements:
Holly Bik
Abstract
Qubit4 Fluorometer Protocol: Invitrogen dsDNA HS Assay Kit (REF 32854); Qubit4 protocol used to test dsDNA concentrations in samples.
Qubit4 Fluorometer Protocol: Invitrogen dsDNA HS Assay Kit (REF 32854)
Set up two assay tubes (Qubit assay tubes REF Q32856) for the standards (Standard #1 and Standard #2) and one assay tube for each sample.
Preparing working solution:
Dilute the Qubit reagent 1:200 in Qubit buffer: 1 µL of Qubit dsDNA HS Reagent *200X (multiplied by number of samples + one extra sample) for every 199 µL of Qubit dsDNA HS Buffer (multiplied by number of samples + one extra sample). Make enough mix for the total number of samples including Standard #1 and Standard #2.
Prepare standards separately (Standard #1 and Standard #2): 190 µL of working solution and 10 µL of standard, to a final volume of 200 µL for each standard.
Tip: Slowly mix up and down with pipette tip.
Prepare samples: 198 µL of working solution and 2 µL of extracted DNA, to a final volume of 200 µL for each sample.
Tip: When pipetting DNA, make sure to mix slowly with pipette tip to prevent shearing. Slowly mix this mixture up and down with pipette tip as well.

Vortex all tubes and centrifuge briefly. Remove any bubbles before inserting tubes into the Qubit4 Fluorometer.

Setting up the Qubit4 Fluorometer: (a) select dsDNA, (b) choose dsDNA HS.
Insert standard tubes into the Qubit Fluorometer and take readings. Note: make sure that standard readings are accurate before moving on to the samples. (Standard #1: low, Standard #2: high). Specify the volume of DNA used and then take readings of your samples.