Boston University 2006:2006-9-07
Thursday 7 September 2006
We grew up M30109 last night and I removed it from the inc at 11:25 am this morning. Next, we need to culture it in LB Amp+Clm broth, miniprep it, and then electroporate it into EnvZ- and WT strains of E. coli.
- we need to make LB stock broth (2 500 ml bottles)
- we need Ampicillin stock
- we need Chloramphenicol stock
Then we can make 5ml of LB Amp+Clm broth in a 5ml tube to culture our E.coli containing M30109.
See this OWW protocol on making cultures for reference when making the broth.
I'll post a little protocol for making antibiotic stocks later.
It would be advantagous to preserve an amount of the M30109 culture (grown from a single colony of E. coli containing M30109 up in LB Amp+Cml broth) for posterity and as a defense against any mistakes we may make in the future. We can do this by making a glycerol stock. Use the Endy lab protocol for now because we have 40% glycerol on hand (the 100% the Quigen protocol calls for is so viscious it's hard to work with).
- The Quigen Glycerol stock protocol uses straight-up 100% glycerol.
- There's also one on OWW from the Endy lab.
We used Tom Knight's electroporation protocol last time and it worked great.