Jun 01, 2024

Public workspaceSubsampling Ethanol Preservative from Zooplankton Museum Collections for DNA Extractions

DOI
dx.doi.org/10.17504/protocols.io.j8nlk888wl5r/v1
  • 1University of British Columbia
Andreas Novotny
Open access
DOI: dx.doi.org/10.17504/protocols.io.j8nlk888wl5r/v1
External link: http://hakai.org
Protocol Citation: Andreas Novotny 2024. Subsampling Ethanol Preservative from Zooplankton Museum Collections for DNA Extractions. protocols.io https://dx.doi.org/10.17504/protocols.io.j8nlk888wl5r/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 31, 2024
Last Modified: June 01, 2024
Protocol Integer ID: 100973
Abstract
This protocol is used to sample DNA from archived samples preserved in ethanol, without having to subsample or split the actual zooplankton biomass.
Guidelines
MIOP: Minimum Information about an Omics Protocol
MIOP TermValue
analysesNucleic Acid Extraction
audiencescientists
broad-scale environmental contextmarine biome ENVO_00000447
creatorAndreas Novotny
environmental mediumsea water [ENVO:00002149]
geographic locationNorth Pacific Ocean [GAZ:00002410]
hasVersion1
issued2017
languageen
licenseCC BY 4.0
local environmental contextcoastal sea water [ENVO: 00002150]
materials requiredSterile workbench, Fume Hood, Centrifuge, Incubator
maturity levelMature
methodology categorySample collection
personnel required1
projectBiomolecular surveys of marine biodiversity in the Northern Salish Sea, BC
publisherHakai Institute, Ocean Observing Program
purposeDNA Extraction
skills requiredsterile technique | pipetting skills
targetDNA
time required1 day
AUTHORS
PREPARED BY All authors known to have contributed to the preparation of this protocol, including those who filled in the template.AFFILIATIONORCID (visit https://orcid.org/ to register)DATE
Andreas NovotnyUniversity of British Columbiahttps://orcid.org/0000-0001-8910-61832024

RELATED PROTOCOLS
PROTOCOL NAME AND LINKISSUER / AUTHORRELEASE / ACCESS DATE
This is a list of other protocols which should be known to users of this protocol. Please include the link to each related protocol.
ACRONYMS AND ABBREVIATIONS
ACRONYM / ABBREVIATIONDEFINITION
GLOSSARY
SPECIALISED TERMDEFINITION

BACKGROUND This protocol is used to sample DNA from archived samples preserved in ethanol, without having to subsample or split the actual zooplankton biomass.
Spatial coverage and environments of relevance As part of the Hakai Institute Ocean Observing Program, biomolecular samples have been collected weekly, from 0 to near bottom (260 m), to genetically characterize plankton communities in the Northern Salish Sea since 2015, developing a climatology from which we can begin uncover the physical, chemical and biological drivers of community and functional change in the dynamic coastal waters of coastal British Columbia.

This protocol has been used as an alternative source of genetic information, when it is not practical to remove biomass from the zooplankton samples.
Personnel Required 1 Technician
Safety Identify hazards associated with the procedure and specify protective equipment and safety training required to safely execute the procedure!
Training requirements Sterile technique, pipetting skills. Work-safe laboratory practices.
Time needed to execute the procedure
1 h
Protocol materials
ReagentSterivex Filter (0.2 um)Merck Millipore (EMD Millipore)Catalog #SVGPL10RC
Step 5
Before start
Read background information, MIOP and BePOP-OBON information under the "Guidelines" tab.
PREPARATIONS
PREPARATIONS
Prepare Sucrose Lysis Buffer (SLB):
Protocol
Sucrose lysis buffer
NAME
Sucrose lysis buffer
CREATED BY
Andreas Novotny

ETHANOL SUBSAMPLING
ETHANOL SUBSAMPLING
To re suspend DNA in the zooplankton sample, invert the sample jar three times.
Let settle for 30 minutes
Use a serological pipette to remove 50 ml of the ethanol preservative, and transfer to a 50 mL falcon tube.
Use a syringe to push the ethanol through a ReagentSterivex Filter (0.2 um)Merck Millipore (EMD Millipore)Catalog #SVGPL10RC .

Seal the outflow of the filter with parafilm.
Add 1800 μL sucrose lysis buffer (SLB).
Seal the inflow opening with parafilm.
Label filter units and store them at -80°C for downstream DNA extraction.
DNA EXTRACTION
DNA EXTRACTION
Follow the same extraction procedures as for Environmental DNA:
Protocol
DNA Extraction from 0.22µm Sterivex Filters - Phenol-Chloroform
NAME
DNA Extraction from 0.22µm Sterivex Filters - Phenol-Chloroform
CREATED BY
Andreas Novotny

Alternative extraction method:

Protocol
DNA Extraction from 0.22µm Sterivex Filters - Qiagen Blood and Tissue Kit
NAME
DNA Extraction from 0.22µm Sterivex Filters - Qiagen Blood and Tissue Kit
CREATED BY
Andreas Novotny