Cryopreservation of Nostoc punctiforme in DMSO                                                                           5/2000

1.  Grow the desired strain in liquid until a fairly dense, but healthy culture is achieved.

2.  Filter sterilize DMSO using sterile polycarbonate filters (other types of filters will dissolve with DMSO).  
     Store the sterilized DMSO at room temperature.

3.  Add 5% DMSO in a sterile manner to the cell culture.  Mix.  We typically freeze 10 x 1 ml for each strain:  
     9.5 ml of culture +0.5 ml DMSO.

4.  Place 1.0 ml of the culture into sterile freezing (cryo-) vials.

5.  Place tubes in -80C freezer.  We have found that quick freezing the culture doesn't increase viability of cells.

6.  To revive a previously frozen culture, simply thaw the tube at room temperature.  As soon as the sample is            
     liquid, pipette the cells into growth medium immediately.  Place the culture under standard growth conditions.