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Finkelstein Lab Wiki

Bacterial Antibiotics

Please make sure to read the notes at the bottom regarding stability and long-term storage of some of these antibiotics.

Working concentrations and stock solutions taken from http://molbiol.edu.ru/eng/protocol/01_05.html (dead link)

Contents

Section titled “Contents”
  1. Stock solutions and working concentrations
  2. Notes
  3. References

1. Stock solutions and working concentrations

Section titled “1. Stock solutions and working concentrations”
Stock Solutions Storage Working Concentrations
Weak Control Strict Control
(mg / ml) ˚C µg/ml dilution µg/ml dilution
bacterial_antibiotics#Ampicillin 50 (H2O) -20 30 0.6 µl/ml 60-125 1.2-2.5 µl/ml
bacterial_antibiotics#Carbenicillin 100 (H2O or 50% EtOH) -20 30 0.3 µl/ml 100 1 µl/ml
bacterial_antibiotics#Chloramphenicol 34 (EtOH or MeOH) -20 10 0.3 µl/ml 170 5 µl/ml
bacterial_antibiotics#D-cycloserine 10 (0.1M NaP pH8.0) +4 2 20 µl/ml
Gentamycin 50 (H2O) -20 15 0.3 µl/ml
Hygromycin B 10 (H2O) -20 10 1 µl/ml 400 40 µl/ml
Kanamicin 50 (H2O) -20 10 0.2 µl/ml 50 1 µl/ml
Kasugamycin 10 (H2O) -20 1000 100 µl/ml
Nalidixic acid 5 (H2O, pH to 11 w/NaOH) -20 15 3 µl/ml
Neomycin 10 (H2O) -20 100 10 µl/ml 800 80 µl/ml
bacterial_antibiotics#Rifampicin 34 (MeOH) -20 150 4.4 µl/ml
Nourseothricin — — — - µl/ml
Spectinomycin* 10 (H2O) -20 100 10 µl/ml
Streptomicin* 100 (H2O) -20 10 0.1 µl/ml 50 0.5 µl/ml
bacterial_antibiotics#Tetracycline 5-12.5 (EtOH or 70% MeOH) -20 12.5 1 µl/ml 50
100		 10 µl/ml
20 µl/ml
Zeomycin -20
G418 200 (H2O) -20 200 1 µl/ml

2. Notes

Section titled “2. Notes”

*Be aware: some competent cells have natural antibiotic resistance. Ex: DH10b cells are Strep/Spect resistant.

2.1 Ampicillin

Section titled “2.1 Ampicillin”
  • Common abbreviation: Amp
  • filter sterilization; or preparation from the sterile water and the sterile powder;
  • store at -20˚C in small quantities (about 10 freezing-thawing cycles);
  • instead of H2O it is possible to use 50% glycerol. The advantage: solution is liquid at -20˚C.
  • some laboratories use 96% (or 50%) EtOH for ampicillin solubilization. There are two advantages in this method:
    • solution is liquid at -20˚C
    • the solution is self sterilizing.
  • But the disadvantage it very serious: ethanol is a weak catalyst of b-lactam ring disruption. So, solution is unstable.

2.2 Carbenicillin

Section titled “2.2 Carbenicillin”
  • Common abbreviation: Carb
  • More expensive than Amp but is also more stable when added to bacteria. This is what we use in the lab.
  • Filter sterilization; or preparation from the sterile water and the sterile powder;
  • store at -20˚C in small quantities (about 10 freezing-thawing cycles);

2.3 Chloramphenicol

Section titled “2.3 Chloramphenicol”
  • Common abbreviation: Cam
  • store dry powder at 4˚C, solution at -20˚C (in EtOH);

2.4 D-cycloserine

Section titled “2.4 D-cycloserine”
  • Unstable solution, prepare just before use

2.5 Rifampicin

Section titled “2.5 Rifampicin”
  • Light sensitive, store in foil

2.6 Tetracycline

Section titled “2.6 Tetracycline”
  • Common abbreviation: Tet
  • Solution in EtOH, store at -20˚C.
  • Light sensitive, store in foil.
  • I’ve seen some references claim that Mg has to be omitted from media for selection because Mg can degrade Tetracycline. Need a ref…

3. References

Section titled “3. References”