Feb 03, 2026
Protocol for environmental Nucleic Acid (RNA and DNA) extraction from filter sample using MagAttract PowerWater DNA/RNA Kit
- Shu-Kuan Wong1,
- Yoko Makabe-Kobayashi2,
- Koji Hamasaki2
- 1National Institute of Polar Research;
- 2Atomosphere and Ocean Research Institute, The University of Tokyo
Protocol Citation: Shu-Kuan Wong, Yoko Makabe-Kobayashi, Koji Hamasaki 2026. Protocol for environmental Nucleic Acid (RNA and DNA) extraction from filter sample using MagAttract PowerWater DNA/RNA Kit. protocols.io https://dx.doi.org/10.17504/protocols.io.ewov11qkpvr2/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: August 20, 2025
Last Modified: February 03, 2026
Protocol Integer ID: 225069
Keywords: DNA extraction, MagAttract, RNA extraction, microbe, water, filter, Sterivex, environmental Nucleic Acid, PowerWater DNA/RNA Kit, eDNA, eRNA, extraction of environmental rna, using magattract powerwater dna, magattract powerwater dna, protocol for environmental nucleic acid, environmental rna, environmental nucleic acid, microbial ena, rna kit, rna kit this protocol, water sample, downstream molecular analysis, extracted ena, extraction from filter sample, rna, filter sample, µm polycarbonate filter, µm sterivex filter unit, dna, polycarbonate filter, extraction, pcr
Abstract
This protocol describes the extraction of environmental RNA and DNA (eNA) from water samples filtered through 0.22 µm Sterivex filter units or 3.0 µm polycarbonate filters using the MagAttract PowerWater DNA/RNA Kit (QIAGEN, 27800-4-EP).
The procedure has been modified for manual handling and employs a Microsmash Beads Beater (MS-100).
The extracted eNA can be used for downstream molecular analyses such as next-generation sequencing (NGS), cDNA library construction, RT-PCR, and qPCR to specifically target microbial eNA.
Guidelines
This protocol has been modified Manual provided by QIAGEN:
-MagAttract®︎PowerWater®︎ DNA/RNA Kit(QIAGEN)
HB-2282-001_1104587_HB_MA_PowerWater_DNARNA_0418_WW.pdf194KB Materials
- Samples
- Sterivex-GP 0.22 μm cartridges (Merck Millipore, SVGP01050) :
For both RNA and DNA samples, cartridges were filled with 1.6 mL of RNAlater‱ Stabilization Solution (Invitrogen‱, AM7020) and stored at −20 °C.
For DNA-only samples, cartridges were stored without RNAlater at −80 °C.
- Nuclpore membranes (PC MB, 47 mm, 3.0 μm) :
For both RNA and DNA samples, membranes were treated with RNAlate‱ Stabilization Solution after that stored at −20 °C.
For DNA-only samples, membranes were stored without RNAlater at −80 °C.
- Reagents
- MagAttract®︎PowerWater®︎ DNA/RNA Kit(384) (QIAGEN, 27800-4-EP)
- RNase AWAY (Thermo Fisher Scientific, 7002)
- 2-Mercaptoethanol (β-ME:beta-mercaptoethanol)（SIGMA, M3148-25ML)
- Ethanol(99.5) for Molecular Biology (FUJIFILM, 054-07225)
- Equipments
- Designated RNA work area and Biological Safety Cabinet
- Microsmash Beads Beater (TOMY, MS-100)
- Microcentrifuge for 5 mL : Centrifugal force needed: 4500 x g
- Digital VORTEX-GENIE 2 (Scientific Industries, SI-A236)
- DynaMag‱-5 Magnet (Thermo Fisher Scientific, 12303D)
- Aluminum block thermostatic bath for 5mL tubes : 60 - 65℃
- Sterile forceps : For transfering the filter into a 5 mL tube
- Pipettes : 1000μL - 100μL
- Consumables
- Filter tips : 1000μL - 100μL
- 5 mL tubes (WATSON 233-150C)
- 1.5 mL tubes
- Terumo Syringes 10mL for Vaccination Slip Tip White (TERUMO, SS-10SZ) : For pushing
out the RNAlater from the Sterivex cartridge
- Parafilm
Troubleshooting
Before start
Wear gloves throughout the extraction process.
Use filter tips to minimize the risk of contaminations.
All steps have to be conducted within a designated RNA work area.
a) Turn on Biological Safety Cabinet blower. Clean the work surfaces with RNase AWAY (Thermo Fisher Scientific, 7002) to remove nucleases. Then Lower the sash and run the UV light for 30 minutes. Switch the UV light back to white light; you are ready to begin.
Before Starting
15m
Warm Solution MBL at 60°C for 15-20 minutes to dissolve precipitates.
60 °C 00:15:00
15m
Warm Clear-Mag RNase-Free Water at 65°C.
65 °C
To extract both DNA and RNA or only RNA, add 25μL of β-mercaptoethanol (β-ME) to 975 μL of Solution MBL.
25 µL
Sample Transfer to Bead Tube
Expel the RNAlater from the Sterivex cartridge using a 10 mL syringe (TERUMO, SS-10SZ).
Cut open the Sterivex cartridge with a tube cutter. Using a surgical scalpel (Akiyama MEDICAL MFG, Replacement blade No. 15) and two sets of sterile forceps, grasp the filter membrane at opposite edges and transfer the filter into a 5 mL PowerWater DNA Bead Tube (provided) with the top side (sample side) facing inward.
Sample Lysis
20m 35s
Add 1.5 mL of pre-warmed Solution MBL to each PowerWater DNA Bead Tube.
1.5 mL
Place the PowerWater DNA Bead Tubes into a Microsmash Beads Beater (TOMY, MS-100). Disrupt the sample at 3,800 rpm for 1 minute, rest for 30 seconds, and then repeat disruption again at 3,800 rpm for 1 minute.
3800 rpm, Room temperature , 00:01:00
00:00:30
3800 rpm, Room temperature , 00:01:00
Equipment
Microsmash Beads Beater
NAME
TOMY
BRAND
MS-100
SKU
LINK
2m 30s
Centrifuge the PowerWater DNA Bead Tubes at 4500 x g for 1 minute at room temperature.4500 x g, Room temperature, 00:01:00
1m
Transfer the supernatant to a clean 5 mL tube (WATSON 233-150C). To recover as much supernantant as possible, insert the pipette tip through the beads to the bottom of the bead tube.
Note: The supernatant may still contain some bio-solid particles.
Add 200 μL of Solution IRS to each tube and seal with Parafilm.
200 µL
Vortex horizontally at 500 rpm for 5 seconds to ensure that the solution is well mixed.
500 rpm, Room temperature , 00:00:05
Equipment
Digital VORTEX-GENIE 2
NAME
Scientific Industries, Inc
BRAND
SI-A236
SKU
LINK
5s
Incubate at room temperature for 5 minutes.
Room temperature 00:05:00
5m
Centrifuge the 5 mL tubes at 4500 x g for 6 minutes at room temperature .
4500 x g, Room temperature, 00:06:00
6m
Avoiding the pellet, transfer the entire volume of supernatant to a new 5 mL tube.
Centrifuge again at 4500x g for 6 minutes to remove any residual particulates that may have carried over.4500 x g, Room temperature, 00:06:00
6m
Transfer all supernatant (~1 mL) to a new 5 mL tube (WATSON 233-150C), carefully avoiding any residual pellet.
<STOPPING POINT> If needed, store the supernatant in the 5ml tube at 4°C for several hours before continuing the protocol.
Binding the Magnetic Beads
23m
Ensure ClearMag Beads are thoroughly resuspended by shaking or rocking the bottle, and re-agitate the beads every 3 minutes.
For each preparation, add 20 μL of ClearMag Beads and 1 mL of ClearMag Binding Solution. Seal the tube with Parafilm.
1 mL 20 µL
Mix the beads and the lysate by placing the tube horizontally on a vortex at the lowest setting (VORTEX-GENIE 2 = 500 rpm) for 20 minutes to allow DNA binding.
500 rpm, Room temperature , 00:20:00
Equipment
Digital VORTEX-GENIE 2
NAME
Scientific Industries, Inc
BRAND
SI-A236
SKU
LINK
20m
Place the tube on a magnetic stand and allow about 3 minutes for the beads to completely adhere to the magnet.
00:03:00
Equipment
DynaMag™-5 Magnet
NAME
Invitrogen™
BRAND
12303D
SKU
LINK
3m
Carefully remove and discard the supernatant without disturbing the beads.
Washing the Magnetic Beads
29m
Add 500 μL of ClearMag Wash Solution and remove the tube from the magnetic stand.
500 µL
Resuspend the beads thoroughly by pipetting up and down for 10 times or by vortexing at the lowest speed for 10 sec, until all beads are fully resuspended.
Place the tube on the magnetic stand and allow 3 minutes for the beads to adhere to the magnet. Carefully remove the supernantant.
00:03:00
Equipment
DynaMag™-5 Magnet
NAME
Invitrogen™
BRAND
12303D
SKU
LINK
3m
Repeat steps 21 - 23 two more times (for a total of 3 washes) using 500 μl of ClearMag Wash Solution each time.
500 µL 00:03:00
6m
Using a 200 μL pipette tip, carefully remove any remaining wash from the tube, taking care not to disturb the beads.
To ensure all wash solution is removed, leave the tubes open in a biosafety hood for 5 minutes, place them on the magnetic stand to remove any remaining liquid, and dry again for 5 mins or until the beads are visibly dry.
00:20:00
20m
Elution of Nucleic Acid
8m
Resuspend the beads in 100 μL (both RNA and DNA) or 50 μl (RNA or DNA only) of Clear-Mag RNAse-free water to elute the nucleic acid.
100 µL
To elute the nucleic acid, heat at 65°C for 5 minutes, flicking the tube occasionally to mix.
65 °C 00:05:00
5m
Place the tube on the magnetic stand and allow 3 minutes for the beads to adhere to the magnet.
00:03:00
Equipment
DynaMag™-5 Magnet
NAME
Invitrogen™
BRAND
12303D
SKU
LINK
3m
Keep the tube on the magnetic stand and carefully open the lid, then transfer the eluate to a new 1.5 mL tube.
The purified nucleic acids can be used immediately. Alternatively, store the DNA and RNA at -80°C for long-term preservation.
Options
For DNA samples: Proceed to the RNase digestion step. To remove RNA contamination, add 2 µL of RNase A (10 mg/mL, Fermentas) to 20 µL of DNA and incubate for 3–4 hours at 37 °C. After enzyme digestion, remove the RNase A using Macherey-Nagel's NucleoSpin gDNA Clean-up Kit, following the manufacturer's protocol.
For RNA samples, proceed to the DNAse digestion step using TURBO DNA-free‱ Kit (Invitrogen, AM1907) and cDNA library preparation steps using PrimeScript‱ II 1st strand cDNA Synthesis Kit (TaKaRa, 6210A) .