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Selective assay for herpes simplex viruses expressing thymidine kinase.

J Campione-Piccardo, W E Rawls, S Bacchetti

Journal of Virology

|August 1, 1979

Summary

A new method efficiently selects herpes simplex viruses with functional thymidine kinase (TK+) from mixed populations. This technique enables precise detection and quantification of TK+ virus fractions, even at low frequencies.

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Area of Science:

Virology
Molecular Biology
Genetics

Background:

Herpes simplex virus (HSV) research often requires distinguishing between viral strains with functional thymidine kinase (TK+) and non-functional (TK-) variants.
Accurate selection and quantification of TK+ HSV are crucial for studying viral genetics, reversion frequencies, and developing antiviral strategies.

Purpose of the Study:

To develop and validate a novel technique for the selective isolation and quantification of TK+ herpes simplex viruses.
To enable the detection of TK+ virus fractions at very low frequencies within predominantly TK- populations.
To facilitate the analysis of TK+ revertants from TK- mutant HSV strains.

Main Methods:

Infection of TK- cells with mixed populations of TK+ and TK- HSV.
Incubation under a liquid overlay medium containing methotrexate to select for TK+ cells.
Addition of fresh uninfected TK- cells at 48 hours post-infection to maintain culture viability.
Detection and quantitation of the TK+ virus fraction using the developed selection method.

Main Results:

The developed technique successfully selected and quantified TK+ herpes simplex viruses from mixed populations.
The method demonstrated sensitivity in detecting TK+ fractions present at frequencies as low as 10(-5).
The technique was effectively applied to estimate reversion frequencies and analyze TK+ revertants from TK- mutant HSV type 1 strains.

Conclusions:

A robust and sensitive method for selecting and quantifying TK+ herpes simplex viruses has been established.
This technique significantly advances the ability to study viral reversion and genetic variation in HSV.
The method provides a valuable tool for herpes simplex virus research, particularly in genetic analysis and mutant strain characterization.

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