Projet Sylvan

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Goal

Etc...


Protocols

Growth Media

LB Agar

More details here.

Protocol To make 500mL of LB agar (makes about 25 LB agar plates):

Weigh out the following into a 500mL Schott Bottle:

  • 5g NaCl
  • 5g Tryptone
  • 2.5g Yeast Extract
  • 7.5g Agar
  • add dH2O to 500mL


Swirl to mix - the contents do not have to be completely in solution, but any powder left on the sides of the flask will caramelize on the glass during autoclaving.

Cover the top of the flask with aluminum foil or with a lid Do not close tightly as it may explode!

Label with autoclave tape (if you have some).


Autoclave on the liquid setting for 20 minutes (be aware that the full autoclave's cycle takes about 3 hours to complete)


After removing the solution from the autoclave (at 75 °C), allow the agar solution to cool to 55 °C.


When pouring plates, keep your bench area sterile by working in a laminar flow hood (turn it on 30 minutes before and clean with ethanol before use).


Pour ~ 15-20mL of LB agar in each 96mm polystyrene Petri dish.


Note: Pour slowly from the flask into the center of the petri dish. When the agar has spread to cover about 2/3 of the dish stop pouring and the agar should spread to cover the entire plate. You may need to tilt the plate slightly to get the agar to spread out completely. If you pour in too much, the plate will be fine, but it will reduce the number of plates you can make per batch.


Do not place the lids on the plates before they are cold (allow them to cool for 30 minutes, until solidified), then invert the plates. Let sit for several more hours or overnight.


Label the bottom of plates with date and store in plastic bags or sealed with parafilm at 4°C.


Light Glucose medium

In 500 mL Schott Bottle:

  • 2 g Glucose
  • 2 g Yeast Extract
  • 1 g NaCl
  • 8 g Agar
  • add dH2O to 500mL

Same procedure as above to make petri agar plates.