Préparation de boite LB agar

1.                  Microwave method for 100 mL agar (about 6 x 10 cm Petrie dishes)

2.                  Use 1 liter Pyrex beaker

3.                  Weigh out LB-agar powder and add sufficient water

a.       2.75 grams per 100 mL water

b.      Mix thoroughly by swirling

4.                  Cover top with plastic wrap (2 layers) and punch 5-10 holes in top to allow venting

5.                  Microwave on high until it just starts boiling (about 1 min)

6.                  Need to boil for at least 1 minute to sterilize

a.       Hit “beverage” key once after it starts to boil

7.                  VERY HOT!!

a.       Use orange hot mitts

b.      Always hold beaker with 2 hands

8.                  Sit on bench until cooled down

a.       Generally 5-10 min

b.      When can touch without getting burnt

9.                  Once cool (not set) add antibiotic of choice

a.       Ampicillin (final conc = 100 ug / mL)

                                                              i.      Add 100 uL of 100 mg/mL sterile solution per 100 mL agar

10.              Mix by swirling

11.              Pour into sterile Petrie dishes

a.       Need enough to cover bottom

b.      About 15-20 mL per regular (10 cm) dish

c.       Apply dish cover with one edge slightly ajar to allow venting

12.              Allow to set and dry out for several hours to overnight at room temperature

13.              Label and store in refrigerator for up to 2 weeks