Growing Bacteria

This pages covers growing bacteria that has optionally been transformed with some genes.

Liquid culture

To grow an overnight liquid culture based on existing liquid culture, put the following into a 100ml Erlenmayer flask:

  • 25ml of liquid LB
  • 25μl of our ampicillin solution, which is a 100μg/ml concentration
  • 200μl of existing overnight liquid culture
  • optional: if you want it to glow, add 125mg of arabinose to the flask, which is 625μl of our stock (20%) arabinose solution, and gives a final concentration of 5mg/ml

Bacteria in agar

To create some agar that has bacteria in it, put the following into each ml of LB with agar:

  • 1μl of our ampicillin solution, which is a 100μg/ml concentration
  • 55μl of existing overnight liquid culture
  • optional: if you want it to glow, add 25μl of our (20%) arabinose solution, which gives a final concentration of 5mg/ml

Use 20ml of LB with agar in a normal petri dish, which is in total:

  • 20ml of LB with agar
  • 20μl of our ampicillin solution
  • 1.1ml of existing overnight liquid culture
  • optional: if you want it to glow, add 500μl of our 20% arabinose solution

First boil the LB agar, than wait until it cools below 42°C, then add all of the above. Make sure to pour it into the dishes before it starts to solidify.

Bacteria on top of agar

Basically use the same agar that is used above, as when putting bacteria into the agar, but of course leave the bacteria out :) After the agar solidifies in the petri dish, make streak the bacteria on it with an innoculation loop.

Bacteria in ‘top agar’

The idea with ‘top agar’ is that it’s not that solid, so that bacteria can spread around in it more easily. So still, the bacteria is inside agar, but in a less solid agar.

Bacteria in top agar is basically two layers of agar: first, create a layer of 14ml per plate as above (without the bacteria in it). Then create an LB agar mixture which is 50-50% LB-agar and liquid LB. This will basically give you an LB-agar mixture with 0.75% agar in it. Put the bacteria into this LB-agar, and spread 6ml of it on top of each plate with the regular agar already in it.

So, for each petri dish, you will have 14ml of LB-agar, consisting of:

  • 14ml of LB with agar
  • 14μl of our ampicillin solution
  • optional: if you want it to glow, add 250μl of our (20%) arabinose solution

and 6ml of ‘top agar’ on top of it, consisting of:

  • 3ml of LB with agar
  • 3ml of liquid LB
  • 3μl of our ampicillin solution
  • 330μl of existing overnight liquid culture
  • optional: if you want it to glow, add 150μl of our (20%) arabinose solution

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