Glycerol stocks (bacterial archive)

Bacterial techniques

Long-term −80 °C storage of any sequence-verified strain. Make a glycerol stock before the first experiment using a new clone — never rely on a plate alone.

You'll need

Saturated overnight culture in LB + antibiotic (16 h, 37 °C, 220 rpm)
50% glycerol, sterile (autoclaved or filter-sterilised)
2 mL screw-cap cryotubes with O-rings (NOT snap-cap — they pop at −80 °C)

Procedure

In a labelled cryotube, mix:
- 500 µL overnight culture
- 500 µL 50% glycerol
Final glycerol concentration: 25%.
Vortex briefly to mix.
Label with: strain, plasmid, antibiotic, your initials, date. Tape over the cap if ink rubs off.
Snap-freeze in liquid nitrogen or place directly into −80 °C in a labelled box.
Log the location (box, row, column) in your strain database.

Recovering a strain

Take the tube out of −80 °C and place on dry ice.
Scrape the surface with a sterile loop or pipette tip — do not thaw the whole stock.
Streak onto a fresh LB+antibiotic plate. Re-freeze immediately at −80 °C.
Incubate 37 °C overnight. Pick a single colony for downstream work.

Naming convention (suggestion)

Pick one and stick with it across the lab. Example:

YC-pYC042-DH5α-2026-04-26-1
(initials – plasmid ID – strain – date – replicate)

Tips

Always store at least 2 independent stocks of irreplaceable strains, in two different freezers if possible.
For very labile strains (e.g., recA⁺ with repeats), drop glycerol to 15% and store at −80 °C only — never −20 °C.
Glycerol stocks are stable for 5+ years if kept at < −70 °C without warm cycles.