TABLE 1.
Primers used for sequential side-directed mutagenesis to generate the phosphomimetic mutant IRF3-5E/Ea
| Protein | Primer | Sequence |
|---|---|---|
| IRF3-1E | S394E (forward) | CCGTGGATCTGCACATCGAGAATAGCCATCCTCTGAGC |
| S394E (reverse) | GCTCAGAGGATGGCTATTCTCGATGTGCAGATCCACGG | |
| IRF3-2E | S396E (forward) | GGATCTGCACATCGAGAATGAACATCCTCTGAGCCTGACG |
| S396E (reverse) | CGTCAGGCTCAGAGGATGTTCATTCTCGATGTGCAGATCC | |
| IRF3-3E | S400E (forward) | CGAGAATGAACATCCTCTGGAACTGACGAGCGACCAGTAC |
| S400E (reverse) | GTACTGGTCGCTCGTCAGTTCCAGAGGATGTTCATTCTCG | |
| IRF3-5E | T402E/S403E (forward) | GAATGAACATCCTCTGGAACTGGAAGAAGACCAGTACAAGGCGTGTCTGC |
| T402E/S403E (reverse) | GCAGACACGCCTTGTACTGGTCTTCTTCCAGTTCCAGAGGATGTTCATTC | |
| IRF3-5E/E | S384E (forward) | CACGTGACGGTGGTGCAAGCGAATTGGAGAATACCGTGGATCTG |
| S384E (reverse) | CAGATCCACGGTATTCTCCAATTCGCTTGCACCACCGTCACGTG |
a
Full-length IRF3 was used as the template for the first reaction to mutate Ser394 to Glu (IRF3-1E). Subsequent Glu substitutions were made using the product from the previous mutagenesis as the template.