Yesterday I streaked out E.coli containing my plasmid onto LB+Amp100 plates from the glycerol stock, and this morning the colonies had irregular shape and they looked bigger than O/N colonies should (incubated for ~15 hrs at 37'C). The glycerol stock is not mine and I know that previously it got thawed a few times. However, I have a plate from about 24 days ago that I've kept the plate at 4'C wrapped with parafilm and it has nice single colonies. So I was wondering what is the best way to make a new glycerol stock? Should I just take a bunch of colonies from the old plate and re-streak them onto a new plate and then make a liquid culture, or can I directly take one single colony from the old plate and grow the culture for the glycerol stock?