Next to Godliness: Bowser Lab Bench Cleanup Protocol

In this lab, we are trying to find unique genetic signatures of organisms in sediment samples and from isolated cells. Since we prep a lot of samples, there’s always the chance that DNA from one experiment could wander into the tubes for another. This is a Bad Thing, since we don’t have any way of telling whether a bit of DNA is supposed to be there or not. And since our detection techniques are sensitive, even a tiny amount of contamination can make a big difference.

The best way to deal with this is:

• a “scorched earth” strategy: destroying DNA on lab surfaces before we begin
• using barrier pipets and other tools to minimize tube-to-tube contamination
• never, ever “borrowing” a piece of lab equipment for use in another area.

Before you begin any experiment in the PCR or primary-purification areas,
you must perform a Bench Cleanup Protocol.

1. Put on a pair of latex gloves.

2. Remove pipets, pipettors, beakers of tubes, boxes of pipet tips, etc. from the bench surface.

3. Wipe the entire bench surface down with Clorox solution, followed by a wipedown with 70% ethanol. Allow to air dry. The bench surface is now free of all DNA.

4. Wet a Kimwipe with a bit of the Clorox solution. Wipe down the bottoms of the beakers and tip boxes, and place them on the bench. Throw out the Kimwipe.

5. Wet down a Kimwipe with some RNAse AWAY solution. Carefully wipe down all surfaces of each of the pipettors. Clean away the solution with a fresh Kimwipe. Change gloves after cleaning if you have any reason to suspect a pipettor is heavily contaminated with DNA (such as the presence of dried globs of something-or-other). Throw out the Kimwipes, and change gloves.

6. Get a waste container. If you are in the PCR areas, use a fresh, clean 800-ml beaker. If you are in the primary purification room, put a new plastic bag inside the round metal waste container.

7. If there is something that you cannot clean this way, such as some powdered reagents, you can use a short-wave UV light. Ask Andrea to show you how.

You are now ready to begin your experiment.

During the experiment:

• If you spill something that is probably DNA-contaminated, do a spot-clean with a bit of Clorox on a Kimwipe. For example: I tend to drip a little bit of extract onto the benchtop during the first part of a sediment extraction. For that reason, I wipe down the tubes before putting them into the water bath, and then I clean off the entire benchtop again. Never be embarrassed to clean anything.
• If you spill or spray something in an unusual place, like the walls, be extra certain to clean it up immediately. The next person will not think to clean there.
• Don’t let the cleaning solutions touch any of your samples. The cleaning solutions destroy DNA—even the DNA you want to keep around.
• When in doubt, change gloves.
• Use the “sterile inside” technique to retrieve tubes and tips from their containers. The outsides of these containers will not be super-clean. These procedures are meant to ensure that the insides of the containers, where the important stuff is, stay uncontaminated.
  • To get a microcentrifuge tube out of a beaker, take the tinfoil top off, and shake several tubes into the inside of the tinfoil cover. Pick out the tubes you need, and shake the rest back into the beaker. This will keep you from smearing contaminants all over the inside of the beaker when you reach in to get a tube.
  • Don’t handle pipet tips with your hands. Open the box, and use only the tip of the pipettor to pick up the tip. You do not have to close the box every time you take out a tip—that would actually be counterproductive—but you should push the box to the side, to minimize accidental contamination by aerosols/drips as you work.
  • When weighing out reagents, such as sodium chloride, do not stick a spatula into the bottle unless it has already been cleaned with RNAse AWAY. Always use a fresh piece of weighing paper or weighboat. You can contaminate your work just as easily with dirty reagents as you can with a dirty pipet.

After you’re done:

Dispose of your garbage. Do not bring the container (beaker or bag) back into the room.

Do a Bench Cleanup Protocol again. Yes, the next person ought to clean up before they start. This is an insurance policy. Finding and eliminating a hidden source of contamination is a royal pain, and we want to avoid it if at all possible.

Back to Molecular Ecology Techniques