- Materials
-
- ~400 ng gDNA in 10 µl nuclease-free water
- Equipment
-
- P10 pipette and tips
- P100 pipette and tips
- Method of achieving 80° C
-
DNA tagmentation
-
Prepare the DNA in nuclease-free water.
- Transfer ~400 ng genomic DNA into a DNA LoBind tube
- Adjust the volume to 10 μl with nuclease-free water
- Mix by flicking the tube to avoid unwanted shearing
- Spin down briefly in a microfuge
-
Cut off Tube 1 (left hand tube, containing FRL)
-
Using a clean, empty pipette tip, pierce the foil of Tube 1. Take care to not disturb the pellet.
-
Add 10 µl of the input DNA to Tube 1.
-
Mix gently by pipetting up and down. Make sure all liquid is collected at the bottom of the tube.
-
Incubate the tube at room temperature for 1 minute and then at 80° C for 1 minute.
-
Adapter attachment
-
Using a clean empty pipette tip, pierce the foil of Tube 2. Take care to not disturb the pellet.
-
Transfer 10 µl of the tagmented DNA from Tube 1 to Tube 2.
-
Mix gently by pipetting up and down. Make sure all liquid is collected at the bottom of the tube.
-
Incubate the reaction for 5 minutes at room temperature.
-
Using a clean empty pipette tip, pierce the foil of Tube 3 (right-hand tube). Take care to not disturb the pellet.
-
Add 65 µl Resuspension Buffer (RTB) into Tube 3. Mix by pipetting up and down, and make sure all liquid is collected at the bottom of the tube.
This can be done during the 5 minute incubation step for the adapter attachment.
Note: This tube might take longer to resuspend. You can assist the process by pipetting the tube contents up and down, and/or incubating at room temperature.