Table 2.
PS mutants tested in cell culture and in vitro
| Virus variant (genetic locus) | Nucleotide locations (see Fig. 2a/b ) | Sequences of the tested mutated PS a (5′–3′) | 50% Tissue culture infectious dose (TCID)/ml b | Replication c | Translation d |
|---|---|---|---|---|---|
| Wild-type | N/A | N/A | 106 | Yes | Yes |
| PS3-M (VP0) | 745–776 | UGGUGUUGUcAGcagc GUUGAUagcACCAUCA | 106 | Yes | Yes |
| PS6-M (VP0) | 1310–1360 | UUcGGAGCuUUUACgAAcCUUCCACAUGUUcUuAUGAAccUGGCUGAAACC | 106 | Yes | Yes |
| PS7-M (VP3) | 1931–1981 | cUgAGACUgUUcCCAAAcUUGAAUGU gUU UGUgAACAGUUAcaguUACUUU | 106 | Yes | Yes |
| PS9-M (VP3) | 2304–2336 | UCUGCCCAACUGGaagUGUaGUaACAUUCCAGA | 0 | Yes | No |
| PS11-M (VP1) | 2645–2673 | UCACUacUcUUcGCcUAcUUcACUGGUGA | 106 | Yes | Yes |
| PS14-M (2B) | 3524–3571 | GCcGCgACcGAGAUUcUuGAUAAcGAU cUcGUCAAGUUCAUAGUGAAA | 104 | Yes | Yes |
| PS18-M (3D) | 6181–6209 | AGCcGGaUAcuccUUcGUcAAAagcGGCU | 105 | Yes | Yes |
| PS21-M (3ʹ UTR) | 7250–7276 | UAAGAcgAAUGaaac GUUcgUCUUUUG | 0 | Yes | Yes |
aThe nucleotides substituted within each mutant PS are in lower case, the loop region of each PS is italicised and the GxU motifs are underlined
bValues are the mean TCID50 values calculated from two independent experiments
cReplication efficiency of each mutant was tested by staining with anti-dsRNA antibody and by quantitative real time PCR
dTranslation efficiency of each mutant was tested by phosphorimaging of in vitro translation of mutants labelled with 35S-methionine