This protocol uses chlorine gas to surface sterilize Arabidopsis thaliana seeds. This is for when you want to perform a more thorough sterilization than an ethanol wash.
The original protocol is attributed to Maud Hinchee (Monsanto), Steven Clough (UI-Urbana-Champaign), and Andrew Bent (UI-Urbana-Champaign).
Purpose: to reduce contamination during experiments
Materials needed: microcentrifuge tubes (1.5 – 2 mL), pencil and paper, BSC/fume hood, container with lid, 250 mL beaker, bleach, concentrated HCl, pipette
Caution: This technique involves chlorine gas and should only be performed by researchers with biosafety training in a laboratory environment. Wear PPE and use proper ventilation at all times.
- Make sure the microcentrifuge tubes your seeds are in are properly labeled. Pencil and paper work the best, as chlorine gas can sometimes wipe Sharpie clean off a surface.
- Place your tubes inside a container that can be closed with a lid.
- Place that container inside a fume hood.
- Make sure the tubes are open. Centrifuge tube lids off.
- Place a 250 mL beaker inside the container with the tubes.
- Put 100 mL of bleach in the beaker.
- Pipette 3 mL of concentrated HCl into the bleach.
- Immediately seal the lid of the container after this addition.
- Let the seeds sit for 3-5 hours.
- Dispose of all waste material properly.