Saltwater BG - 11 recipe
Bring final volume to 1L .
Adjust pH of the final media to 8.2 .
* Be sure to add K2HPO4 first , then NaSiO3 , then add the others stock solutions to prevent precipitation .
0.2 um filter sterilize into clean , autoclaved containers in a biosafety hood .
Do not autoclave the media as the silica will form precipitate and be cloudy !
To Prepare Conjugation Plates ( 1 / 2 BG - 11 , 5 % LB , 1 % agar ) : Dilute Saltwater BG - 11 1 : 1 with ddH2O .
Add 50mL of 1x LB for each litre of Conjugation Plate media being made .
Add 5g of Bacto Agar to a 1L Earlenmeyer flask for each 500mL of Conjugation Plate media being made .
Distribute 500 mL of Conjugation Plate media to each 1L Earlenmeyer flask .
Cover with flask opening with aluminum foil and autoclave at 121C for 30 minutes .
Allow media to cool while stirring on magnetic stir plate .
Make sure the stirring is not too vigorous as to cause bubbles .
It should cool enough to be able to touch , but not have the agar solidified .
Pour plates and allow to cool overnight .
To Prepare Selection Plates ( 1 / 2 BG - 11 , 1 % agar , + antibiotics ) : Dilute Saltwater BG - 11 1 : 1 with ddH2O .
Add 5g of Bacto Agar to to a 1L Earlenmeyer flask for each 500mL of Selection Plate media being made .
Distribute 500 mL of Selection Plate media to each 1L Earlenmeyer flask .
Cover flask opening with aluminum foil and autoclave at 121C for 30 minutes .
Allow media to cool while stirring on magnetic stir plate .
Make sure the stirring is not too vigorous as to cause bubbles .
It should cool enough to be able to touch , but not have the agar solidified .
Add 500ul of the required antibiotics to the Selection Plate media made .
We make 500uL aliquots of 1000x antibiotic stocks .
It is very important that the media is cool before adding the antibiotics or the heat will destroy them .
Pour plates and allow to cool overnight .
