question on making chemical and electrocompetent cells - (Jul/01/2011 )
I have 2 questions about making competent cells
1) What is the purposed of incubating the cells on ice before harvesting? It is usually included in protocols and there is one that suggest up to 2h "rest" for the cells on ice!
2) What should the final OD for competent cell be? One protocol I've read suggest OD600~375 for electrocompetent cells. What about chemical competent cells? Or is there a rule of thumb how many folds the culture should be concentrated down?
To answer your 2nd Qstn , i would suggest you to harvest cells for preparing chem comptnt at O.D 0.5 or not more than 0.6 (600 nm )....and i usually scale down 10 to 1 ....hope this helps....
YOu should read the original papers on these - Hanahan, 1984 is a good start.