So, you want to isolate that plastic degrading, or bioluminescent, or plasmid-bearing microbe from the rest of a mixed sample. Or maybe you want to grow up a colony from a single bacteria for a clean PCR sample, or to inoculate a liquid culture. To do this you will need to streak a plate! There are many techniques for this, but there are really two key ideas:
- Work cleanly; don’t contaminate your sample. Use an open flame
- You want to thin out the amount of bacteria on your loop
My technique for this is to heat my loop up to orange-hot, then cool it in the agar of the sample I am taking the colony from, being careful not to touch any of the colonies on the plate. Once quenched (So I don’t heat-kill the bacteria), I rub it on the target colony or area on the sample plate. Then I close the sample plate and open the target plate. I like to do 3 streaks. The first one is to spread out the bacteria I picked up on the top 1/3 of the plate. Once this is done, I sometimes flame and cool the loop in the target agar, although it is ok to skip this step. Either way, I then make another zig-zag, starting in the area that I spread the bacteria in, and then moving out onto an unused third of agar. The last streak takes up the last third, and starts in the last area I covered.
If everything works out, you should have isolated colonies like in the picture above. A few common pitfalls are:
- TOO MUCH bacteria. Bacteria are very small, and if you pick up too many, you will end up with a bacterial “lawn”, which is useless for isolating colonies.
- Accidentally killing your sample bacteria by sticking your red-hot loop into them (doh!).
- Streaking back into an already streaked area. This defeats the purpose of streaking, which is to spread out the bacteria. If you go back from a low concentration area to a high concentration area, and then back to the low concentration area, you risk bringing extra concentrated bacteria that would form a lawn into an area where you want single colonies.