Tetracycline

Mode of Action

Tetracyclines (there are many) enter the cell by diffusion through OmpF porins. It inhibits growth by preventing codon-anticodon interactions during translation.

Resistance Mechanism

There are a number of resistance mechanisms, most require tetracycline antiporters. The most commonly used is the tetA(C) gene. The gene product of tetA(C) forms a pore in the inner membrane and exports one tet molecule for every proton imported. Molecular Cloning states that tet resistance has a number of adverse affects on the cell, chief among those being a reduction in growth rate and viability.

Working concentration and Stock Solutions

• Using a plasmid with copy number 15-20, 20μg/ml tet has proved a successful selector. The reduction in growth rate, described above was significant.
• The RajBhandary lab uses 5μg/ml and see a minimal reduction in growth rate and never lose plasmid from their cells.
• Stock solution is 15 mg/ml in 50% ethanol. Add 1 ml per liter of cooled broth or agar. Solutions and plates are light sensitive.
• Tetracycline is incompatible with (chelated by) magnesium ions, and thus unsuitable for use in minimal media

Counterselection techniques

• Fusaric acid can be used to select against tetracycline resistant colonies. See PMID 6259126, and modifications PMID 7007341.
• Nickel ions can also be used for tetR counterselection. See PMID 8954882.

Properties

• Molecular Weight - 480.9 g/mol (Hydrochloride salt), 444.4 g/mol (free base)

References

• Molecular Cloning
• Wikipedia Tetracycline page