Hello. I have two doubts:
1) I am checking protocols for preparing SOB medium. Some of them (including Sambrook et al 2003) suggest autoclaving the broth prior to adding MgCl2 or MgSO4. Some other (more recent) protocols say to mix everything and THEN autuclaving after full dilution and pH adjustment. Wyhy is autoclaving MgCl2 or MgSO4 avised against?
2) Some protocols include MgSO4, some other don't. Is there any major difference in the usefulnes/performance/stability/any other feature of SOB broth with, Vs. without MgSO4?
Thank you!
Dear Rony,
Autoclaving before or after dilution is the same. No differences are expected. All protocols use MgSO4 alone or in combination with MgCl2.
For bacterial growth you need MgSO4.
The following is the recommended protocol:
http://2011.igem.org/Team:Uppsala-Sweden/Project/Protocol
SOB Medium. Used in growing bacteria for preparing chemically competent cells.
Ingredients
0.5% (w/v) yeast extract
2% (w/v) tryptone
10 mM NaCl
2.5 mM KCl
20 mM MgSO4
Per liter:
5 g yeast extract
20 g tryptone
0.584 g NaCl
0.186 g KCl
2.4 g MgSO4
Note: Some formulations of SOB use 10 mM MgCl2 and 10 mM MgSO4 instead of 20 mM MgSO4.
Adjust to pH 7.5. This requires approximately 25 ml of 1M NaOH per liter. Autoclave the bottled SOB
SOC medium is SOB medium with 20 mM glucose. You can do that by adding 20 ml of 20% glucose into 1 liter of SOB. Make sure everything is sterile though.
Hoping this will be helpful,
Rafik
1. MgSO4 is more popular as Cl from MgCl2 (in addition to KCl and NaCl) is increasing the salt concentration undesirably.
2. Always filter sterilize MgSO4 (it is NOT volatile) and add after autoclaving to avoid precipitation and also MgSO4 addition would not effect pH changes.
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