Glycerol Stocks - Preparation of Glycerol Stocks (May/26/2011 )

Hi everyone,

So basically I made a glycerol stock and forgot to put it into the -80 C freezer. I realised this after 2 days at room temperature and put it immediately in the -80 C freezer. Would this affect the viability of cells, because being at RT will only increase their density.

Thanks!

Saeed

Yes, it will decrease the viability. The cells will deplete the nutrients in the tube well before 48 hours are up.

bob1 on Sat May 28 00:08:38 2011 said:

Yes, but will I be able to use it as a glycerol stock to transform? If not, what are the chances of an unwanted mutation if I use 1 ul of my purified plasmid in the cell line and make fresh glycerol stocks ?

Regards,

Saeed

I would not use those cells anymore to make a transformation.
2 days at room temp is long.. most cells might be dead.
Mutations, I dont think there will be more mutations then when with regular growth.
Just make new cells... or are they so special?

Maybe you can just plate them out and make a new stock out of them? By doing this, you also immediatly see if they are still viable.. (plate out a few and if you see they are still viable, its worth a change using them. But to be honest: I would just use new cells, not worth losing a few days trying it with old cells.. but if you have some spare time you can try it)

Thank you.