Question making competent bacterial cells - (Aug/28/2007 )
I wonder if I have to use overnight grown culture before I grow them in a large volume.
Can I just use frozen bacteria in a large LB culture to get OD600 of 0.4?
Thank you for your answer in advance!
Yes you could do that, just make sure that the bacteria are clonal (i.e. you have grown the culture from a single clone) and that you use a sufficient amount of bacteria, otherwise it could take a long time to grow. A good approach is to make a cell bank - grow the cells to mid log phase (OD=0.5 or so) then freeze 1-2ml aliquots in glycerol at -80°C. When you want to grow up a culture for competent cells, just innoculate with one of your aliquots (use a 1% innoculum, e.g. 1ml in a 100 ml culture) and you should get an OD of 0.4 or so in a few hours, depending on the growth conditions.
Great! Thanks a lot!