Name:

Relative freq MxA-, 54 hpi, replicate1

Category:

Growth/Fitness

Property:

Frequency of Occurrence

Units:

unitless

Measurement Technique:

Illumina Sequencing , Viral Competition

Buffers:

Temperature:

pH:

None

Protein Concentration:

Wavelength:

Ionic Strength:

Details:

To confirm that the decreased MxA resistance of polymerase activity correlated with the effect of MxA on viral replication, we carried out competition experiments between wildtype and mutant viruses. Such competition experiments provide a sensitive and internally controlled way to measure the relative fitness of two viral variants. We used reverse genetics to generate influenza viruses carrying wildtype NP, the D51Dsyn mutation, or the D51N mutation. We mixed each mutant virus with wildtype virus at a 1:1 ratio of infectious particles, and infected MxA-expressing or non-expressing cells at a low MOI. At 10 and 54 hours post-infection, we isolated viral RNA and determined the frequency of each variant by Illumina deep sequencing. Experiments were performed in duplicate (replicates 1 and 2). Relative frequencies (mutant/WT) were determined for each of the conditions (MxA+, MxA-) at each time point for each replicate.