# DNA extraction

[EcoEvoDevoLab](https://hackmd.io/@EcoEvoDevoLab/AngeliniLab)

Updated 24 September 2021


## Qiagen columns

If you're using the [Qiagen DNeasy Blood and Tissue Kit](https://www.qiagen.com/us/products/discovery-and-translational-research/dna-rna-purification/dna-purification/genomic-dna/dneasy-blood-and-tissue-kit/) columns for DNA extraction, please just follow the [protocol](https://www.qiagen.com/us/resources/download.aspx?id=68f29296-5a9f-40fa-8b3d-1c148d0b3030&lang=en) with the kit!

## Maxwell 16

Use the Maxwell 16 and the Maxwell 16 Tissue DNA Purification Kit (Promega Cat.# AS1030) for automated extraction of large numbers of samples. The protocol below is based on the instructions from the kit, and works best with relatively large amounts of tissue. Use up to 50 mg of tissue in this protocol. Once the cartridges are unsealed, they are suseptible to contamination, so don’t pause for long periods between steps.

- Place each cartridge to be used into the holder with the ridged side of the cartridge facing toward the numbered side of the rack.
- Carefully peel back the seals from the top of the cartridges. - Remove any residue.
- Place one plunger into well #7 of each cartridge. (Well #7 is the well closest to the ridged side of the cartridge.)
- Transfer your sample into well #1. (Well #1 is the well closest to the cartridge label and furthest from the user.)
- Using a plunger, gently crush the tissue into the lysis buffer already present in the well.
- Turn on the Maxwell instrument.
- Verify that the instrument settings indicate an `SEV` hardware configuration and `Rsch` operational mode. If it's not, use the set-up menu to switch from `LEV` to `SEV` and replace the hardware (the magnetic rods and sample tray).
- Transfer cartridges containing samples and plungers from the cartridge preparation rack onto the instrument’s platform. Ensure that the cartridges are placed into the instrument with the ridged side of the cartridge closest to the door. If you are processing fewer than 16 samples, center the cartridges on the rack.
- Place one blue elution tube for each cartridge into the elution tube slots at the front of the platform.
- Add 300 μl of nuclease-free water to the bottom of each Elution Tube.
- Select `Run` on the Menu screen, and press the `Run/Stop` button to select the method.
- Select `DNA`; then select `Tissue DNA` on the Menu screen. - Next select `OK` at the Verification screen.
- Open the door when prompted. Press the `Run/Stop` button to extend the platform.
- Press the `Run/Stop` button. The platform will retract.
- Close the door. Maxwell will then isolate the DNA!
- Check the timer to see when the run will finish.
- While the machine is running, label 0.5-ml storage tubes “genomic DNA” with the species, population, sex, genotype, and/or tissue of origin and your initials and today’s date.
- When the run ends, the screen will prompt you to press the `Run/Stop` button to extend the platform.
- Remove the elution tubes from the heated elution tube slots, and place them into the Magnetic Elution Tube Rack.
- Power down the machine.
- Transfer the eluted samples into storage tubes by pipetting. Avoid any paramagnetic particles (an insoluble gray powder).
- Discard the blue elution tubes after transfer of the eluted sample. Remove cartridges and plungers from the instrument platform and discard.
- Use the NanoDrop spec to measure the concentration of the DNA. Record this information in your notebook and on the side of the tube. Also note the purity of the sample. Pure DNA should have an A260/A280 ratio > 1.8.
- Store DNA at -20˚C.

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[EcoEvoDevo Lab](https://hackmd.io/@EcoEvoDevoLab/AngeliniLab)