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Antibiotics Used in Molecular Biology

Posted in: DNA / RNA Manipulation and Analysis

Antibiotics are used in a wide range of techniques in molecular biology including molecular cloning and are important for treating pesky mycoplasma contamination in cell cultures. They can also be used to maximize your plasmid yields by reducing protein synthesis, in certain circumstances.

The aim of this post is to provide an easy reference to some of the main antibiotics that are used in molecular biology, their mechanisms, range, and working concentrations. I hope you will find it useful.

Your Personal Antibiotics Reference Guide





Resistance conferred by

Conc. (μg/ml)


Beta-lactam (Penicillins)

Gm+, Gm-

Inhibits transpepsidase required for cell wall synthesis

Beta-lactamase: cleaves the beta-lactam ring of ampicillin


Amphotericin B

Antifungal (Polyenes)

Ye, Fu, My

Binding to ergosterol at the membrane, inducing pore formation and ergosterol sequestration, induction of oxidative damage

Pleiotropic mechanisms: altered membrane composition, ATP-binding cassette transporters, and upregulated thiol metabolic pathway



Beta-lactam (Penicillins)

Gm+, Gm-

Similar to Ampicillin: inhibits transpeptidase required for cell wall synthesis

As for ampicillin: but carbenicillin is broken down more slowly by beta-lactamase



Synthetic (Fluoroquinolones)

Gm+, Gm-, My

Inhibits bacterial DNA gyrase and topoisomerase IV

Pleiotropic mechanisms: genetic mutation of the A subunit of DNA gyrase, and by alteration of drug permeation through the outer membrane of the cell




Gm+, Gm-

Binds to ribosomal 50S subunit, preventing peptidyl transferase required for translation

Chloramphenicol acetyltransferase: adds an acetyl group from ACoA to chloramphenicol, which inactivates it




Gm+, My

Similar to Chloramphenicol

ermC methyltransferase: methylates the 23S rRNA, preventing erythromycin binding to the ribosome




Gm+, Gm-, My

Binds to the 30S ribosomal subunit, blocking the initiation complex and causing frame-shift mutations and inhibition of translation

Kanamycin phosphotransferase: affects the ATP dependent phosphorylation of hydroxyl residues on kanamycin




Gm+, Gm-

Similar to kanamycin: inhibits bacterial protein synthesis through irreversible binding to the 30S ribosomal subunit

Gentamicin acetyltransferase: mechanism similar to chloramphenicol acetyltransferase




Gm+, Gm-

Similar to kanamycin: inhibits bacterial protein synthesis through irreversible binding to the 30S ribosomal subunit

Neomycin phosphotransferase: mechanism similar to kanamycin phosphotransferase



Antifungal (Polyenes)

Ye, Fu

Similar to amphotericin B: formation of a membrane-spanning pore in the fungal plasma membrane

Genetic mutation: resulting in changes in sterol spectrum specificity


Rifampicin (rifampin)

Semi-synthetic (Rifamycins)

Gm+, Gm-

Inhibits DNA dependent RNA polymerase, preventing transcription

Genetic mutation: of the rpoB gene encoding the beta subunit of RNA polymerase that alter residues of the rifampicin binding site on RNA polymerase, resulting in decreased affinity for rifampicin




Gm+, Gm-

Binds to 16S ribosomal subunit, preventing initiation of translation

Streptomycin 3'-adenyltransferase: transfers the adenyl group from ATP onto streptomycin




Gm+, Gm-

Prevents aminoacyl tRNA from binding to 30S subunit

Pleiotropic mechanisms: Tetracycline efflux, ribosome protection, enzymatic inactivation of Tetracycline


Abbreviations: Fu = fungus; Gm(+/-) = Gram positive/negative; My= mycoplasma; Ye = yeast.

This is by no means an exhaustive list of all antibiotics used in molecular biology, so if I have missed out on an antibiotic that you use routinely in your work, please leave a comment and I will add it to the table.

It’s important that you store your antibiotics appropriately, so you can ensure that they are working correctly when you come to use them. For a guide on proper storage and use of antibiotics, check out our related article Antibiotic Stability: Keep Your (Gun)powder Dry. This article also provides information on the best solvent to use for your stock and whether the antibiotic is light-sensitive or not.

When using antibiotics in plates, do you know how long in advance you can prepare them, or if the plates that have been in your cold rooms for the past few weeks are still usable? You might be surprised when you find the answers in our article: Ye Olde Antibiotic Plates.

If you are planning to use ampicillin for selection (for example when cloning or performing protein purification in E. coli), there are several limitations you need to be aware of that might affect how you use this antibiotic. We go into the details of these limitations as well as precautions you can take in our article What’s The Problem With Ampicillin Selection?

Our downloadable wall chart aims to provide an easy reference to some of the main antibiotics used in molecular biology, their mechanisms, range, and working concentrations. Print it off and post it in your lab for some eye-catching yet educational decoration.

Originally published on October 2, 2007.  Updated and revised on 10 December 2019.

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Image Credit: yothinsanchai


  1. Luke on August 16, 2016 at 1:12 am

    Thank you for this table! We use Ciprofloxacin a lot in our lab, would you be able to add this when you have a chance please? Thank you!

  2. Anon on August 4, 2016 at 2:33 pm

    Small correction, amphotericin is now thought to be fungicidal independent of its ability to form a pore by instead “sponging” up ergosterol from the membrane (see Martin Burke lab for details)

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