How to Properly Streak a Single Bacterial Colony

Bacteria are the workhorses of many molecular biology laboratories, and mastering the basic techniques to manipulate bacteria is an important stepping-stone towards achieving great results. When isolating DNA from bacteria, it is important to start with a single colony to ensure a homogenous population of bacteria in your culture. Isolating a single bacterial colony from an agar plate can be more art than science sometimes, but follow these instructions to streak a plate like a pro!

Day 1:

1. Grab your plate from the fridge. Try to remember if the red stripe on the side means Kanamycin or Ampicillin. Give up and grab one of each just to be sure.

2. Squint to determine whether that black spot is mold or a stray Sharpie mark. Stray Sharpie. Definitely. Maybe.

3. Warm plates up in the 37°C incubator for anywhere between a) as long as you can stand, and b) how long it takes you to remember you were warming up plates. If b) exceeds 2 days and your plates have dried out and cracked, gather new plates and repeat from Step 2.

4. Sterilize your work area with ethanol and light your Bunsen burner. Have a half-second panic attack that you’re going to light your bench on fire as you light it.

5. Sterilize your inoculating loop in your Bunsen burner. Do it again just to be sure.

6. Pick up your sample with the loop and hope that it doesn’t sizzle.

7. Deposit sample in first quadrant. Wonder if you’re actually putting any bacteria down at all! If you’re unsure what this means, check out Diagram 1 for a visual guide on how to streak.

8. Sterilize loop. Wait impatiently for it to cool. Streak second quadrant from first quadrant. Is that too big? Too small? Too late now.

9. Repeat Step 8 for third and fourth quadrant, doing the first 2 swipes through the previous quadrant. Wonder whether you really picked up anything.

10. Put plate in incubator agar side up, and incubate overnight. Maintain a persistent low-level anxiety for 12-16 hours about whether or not it’ll work.

Diagram 1: How to streak a plate. The red lines represent the most recently streaked area during the process. Figure by Lynnea Waters.

Day 2:

11. At this point, your bacteria are Schrödinger’s cat—simultaneously growing and not growing in your incubator. Put off checking plates until mentally ready to deal with both outcomes.

12. Congratulations, you have successfully streaked a plate! Note: We can’t guarantee that you will actually get individual colonies, only that you have streaked a plate!