TABLE 2.
Primers used for amplification, cloning, sequencing, and mutagenesis protocols
| Purpose and code | Tmc (°C) | Regiona | Positionb | Sequence | Reference |
|---|---|---|---|---|---|
| Amplification, cloning, and sequencing | |||||
| IRES1 | 62 | 5′ UTR (GI) | 28 | GAGYCCCTCTTGGAAGTC | This study |
| IRES2 | 58 | 5′ UTR (GI) | 869 | AAAATAAGAAGCACCAGTCAC | This study |
| IRES3 | 67 | 5′ UTR (GII-III) | 63 | ATACCTCACCGCCGTTTGC | This study |
| IRES4 | 65 | 5′ UTR (GII-III) | 785 | GTCAAGRCCACTCCCAACAG | This study |
| +697 | 64 | 5′ UTR | 697 | TCAGGGGCATTTAGGTTTTTC | This study |
| 5′ Bgl2 | 75 | 5′ UTR | 63 | CCCAGATCTATACCTCACCGCCGTTTGC | This study |
| 3′ NcoG1RW | 72 | 5′ UTR | 738 | CCCCCATGGTCATTATTATTTAAGAATGAGGAAAAAC | This study |
| 3′ NcoG1F1 | 77 | 5′ UTR | 738 | CCCCCATGGTCATTGTTATTTGAGAATGAGGAAAAAC | This study |
| 3′ NcoG1F2 | 75 | 5′ UTR | 738 | CCCCCATGGTCATTATTGTTTAAGAATGAGGAAAAAC | This study |
| 3′ NcoG1B1 | 77 | 5′ UTR | 738 | CCCCCATGGTCATTATTGTTTGACAATGAGGAAAAAC | This study |
| 3′ NcoG1BF | 72 | 5′ UTR | 738 | CCCCCATGGTCATTATTATTTAAGAATGAGGAAAAAC | This study |
| 3′ NcoG2B1 | 75 | 5′ UTR | 738 | CCCCCATGGTCATAATTATTTGAGAGTGAGGAAAAAC | This study |
| 3′ NcoG3B1 | 75 | 5′ UTR | 738 | CCCCCATGGTCATAATCATTTATTGATGAGGAAAAAC | This study |
| 3′ NcoG3F1 | 71 | 5′ UTR | 738 | CCCCCATGGTCATAATTATTTATAGATGAGGAAAAAC | This study |
| Strain genotyping | |||||
| +2870 | 62 | VP1-2B | 2870 | GACAGATTCTACATTTGGATTGG | 4 |
| −3381 | 64 | VP1-2B | 3381 | CCATTTCAAGAGTCCACACACT | 4 |
| +2870G3 | 62 | VP1-2B | 2870 | AACAGATTCAACTTTTGGACTTG | This study |
| −3368G3 | 64 | VP1-2B | 3368 | CCACACACTCCTACCAGCAG | This study |
| 5′ UTR mutagenesis | |||||
| a324g | 5′ UTR | GAGTCTAAATTGGGGACGCAGATGTTTGGAACGTC | This study | ||
| a324g-R | 5′ UTR | GACGTTCCAAACATCTGCGTCCCCAATTTAGACTC | This study | ||
| t372c | 5′ UTR | AGTGTTAACTTGGCTTTCATGAACCTCTTTGATCTTCCACAAG | This study | ||
| t372c-R | 5′ UTR | CTTGTGGAAGATCAAAGAGGTTCATGAAAGCCAAGTTAACACT | This study | ||
| t255c | 5′ UTR | CTTGCCCTAGGCCCTGGCCGTTGCG | This study | ||
| t255c-R | 5′ UTR | CGCAACGGCCAGGGCCTAGGGCAAG | This study | ||
| t278c | 5′ UTR | CGCCCGGCGGGGCCAACTCCATGAT | This study | ||
| t278c-R | 5′ UTR | ATCATGGAGTTGGCCCCGCCGGGCG | This study |
a
Genotype specificity is indicated in parentheses (GI, GII, and GIII, genotype I, II, or III, respectively).
b
Position of the first 5′ nucleotide on the reference sequence HM-175 (GenBank accession number M14707).
c
Tm, melting temperature.