TABLE 1.
Plasmids and primers
| Plasmid | Siteb | Insert (nt) | Primera |
|---|---|---|---|
| pFlag-Q2779 | nsp4-5 | 8569-9069 | 5′-ATCCACTCGTAAGCTGTATGATGGC |
| 5′-CGGCTGCAGTTATTGAGTTGTAACTTCAAACTC | |||
| pFlag-Q2779N | 5′-AGTAGATTAAACTCTGGTTTTAAG | ||
| pFlag-Q2779E | 5′-AGTAGATTAGAATCTGGTTTTAAG | ||
| pFlag-Q3086 | nsp5-6 | 9490-9981 | 5′-ATCCGCAATTATTAGTGTTAAGGAG |
| 5′-CGGCTGCAGTTAAGTATCCATATATGCCATAAC | |||
| pFlag-Q3086N | 5′-GGTGTTAGATTAAACTCTTCTTTT | ||
| pFlag-Q3086E | 5′-GGTGTTAGATTAGAATCTTCTTTT | ||
| pFlag-Q3379 | nsp6-7 | 10531-10860 | 5′-ATCCGTTTCAGTAGATCAATATAGG |
| 5′-CGGCTGCAGTTAAGTAGAATCTATACAAAATAG | |||
| pFlag-Q3379N | 5′-GCTACAGTTAACGCTAAATTGAGT | ||
| pFlag-Q3379E | 5′-GCTACAGTTGAAGCTAAATTGAGT | ||
| pFlag-Q3462 | nsp7-8 | 10666-11544 | 5′-ATCCGCTAAATTGAGTGATGTAAAG |
| 5′-CGCTGCAGTTATTGCAAAACAACATCAACCTT | |||
| pFlag-Q3462N | 5′-ACTGTATTAAACTCGGTTACTCAA | ||
| pFlag-Q3462E | 5′-ACTGTATTAGAATCGGTTACTCAA | ||
| pFlag-Q3672 | nsp8-9 | 10915-11877 | 5′-ATCCTCGGTTACTCAAGAATTCTCA |
| 5′-CGCTGCAGTTACTGTAAGACAACAACATTAGA | |||
| pFlag-Q3672N | 5′-GTTTTGAACAATAATGAGCTTATG | ||
| pFlag-Q3672E | 5′-GTTTTGGAAAATAATGAGCTTATG | ||
| pFlag-Q3783 | nsp9-10 | 11590-12120 | 5′-ATCCGCAGGTGTAGATCAAGCACAT |
| 5′-CGGCTGCAGTTAGTGTGCTCTACAATAGAGACA | |||
| pFlag-Q3783N | 5′-GTTGTCTTAAACTCTAAAGGGCAT | ||
| pFlag-Q3783E | 5′-GTTGTCTTAGAGTCTAAAGGGCAT | ||
| pFlag-Q4868 | nsp12-13 | 14979-15479 | 5′-ATCCTTTGTTCTCCTTGCATATATC |
| 5′-CGGCTGCAGTTAAATGGACCAATTAGTAGTAGC | |||
| pFlag-Q4868N | 5′-ACGACTTTAAACTCTTGTGGCGTT | ||
| pFlag-Q4868E | 5′-ACGACTTTAGAATCTTGTGGCGTT | ||
| pFlag-Q5468 | nsp13-14 | 16785-17159 | 5′-ATCCGCAGATTCGCAGCATGCACTG |
| 5′-CGGCTGCAGTTAATCACGTGTTATAAACATGTT | |||
| pFlag-Q5468N | 5′-GAAACAAGTCTGAACGGTACAGGT | ||
| pFlag-Q5468E | 5′-GAAACAAGTCTGGAAGGTACAGGT | ||
| pFlag-Q5989 | nsp14-15 | 18219-18719 | 5′-ATCCAGCTTCCGCAATTTGAAAGCT |
| 5′-CGGCTGCAGTTATAAACCTTTCAAAATACGGTT | |||
| pFlag-Q5989N | 5′-TCAGCTCTCAACTCTATCGACAAT | ||
| pFlag-Q5989E | 5′-TCAGCTCTCGAGTCTATCGACAAT | ||
| pFlag-Q6327 | nsp15-16 | 19299-19799 | 5′-ATCCTTTGTATTGTCGGACAATGGT |
| 5′-CGGCTGCAGTTAATCATTATCGACAAGGAGTGT | |||
| pFlag-Q6327N | 5′-CCACAGCTTAACTCAGCATGGACG | ||
| pFlag-Q6327E | 5′-CCACAGCTTGAATCAGCATGGACG | ||
| pFlag-Q6327H | 5′-CCACAGCTTCACTCAGCATGGACG | ||
| pFlag-Q6327P | 5′-CCACAGCTTCCATCAGCATGGACG |
a
Sequences are based on the IBV strain Beaudette P65 (DQ001339.1). The underlined type indicates a mutated codon. Only the forward primer for each mutant is shown, and the reverse primer is completely complementary with it.
b
The location of IBV 3CLpro cleavage sites.