When freezing an overnight exponential or stationary phase bacterial culture in glycerol and growth medium (say 50:50 of a 30% glycerol stock), can you immediately re-use the thawed sample or does it need to be re-streaked and grown again? I appreciate that the common method is to scrape some of the frozen sample and re-streak but in the interests of time and convenience I am considering attempting to test the viability by merely thawing, spinning and re-suspending.
Hi David,
First, remember to never let your frozen glycerol stock thaw out, otherwise you could end harming whatever you have cultured (it's a big shock for the cells). Just be sure to use it quickly and put it back at -80. Secondly (if I understand your question correctly), if you just made a freezer stock of bacteria and wish to use it right now instead of growing it on a plate first and using a colony from there, that is fine. A lot of times this is done for purity reasons but if you feel that they are pure and free of contaminates, go ahead. Why are you planning on spinning them down? For viability checks, I've used freezer stocks and just inoculated a liquid culture from the ice.
I would streak them out every now and then just to be sure they have not gotten contaminated.
Hi!
Don't thaw your glycerol stock, merely dip a (sterile) pipette tip in it and put it in 2 mL LB medium (or equivalent) pre-culture. Then after a few hours either transfer it to a larger volume for midi/maxy prep. I used to start the pre-culture in the morning, and then transfer at the end of the day to 150 mL LB medium. Or inoculate your LB medium for o/n growth at the end of the day (for miniprep volumes). So no need to thaw (bad for your bacterial cells!), centrifuging or resuspending.
David seems to have hit the nail on the head. I've sometimes had to go right to a broth culture in the interest of time, but you obviously lose the visual check for contamination and odd colony morphology. If I do need to go from glycerol right to broth, to be safe, I usually plate a small amount after an hours growth to ensure the glycerol stock wasn't contaminated (especially if it's shared) and the colonies look "correct".
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