Table 1.
Primers used for generating pBAC-C-KCE, donor plasmid pRThGA and identification of the pBAC-C-KCE-E.
| Purpose and Primer | Sequence (5’→3’) | Sequence Designation, Restriction Enzyme Site and Introduction Sequence |
|---|---|---|
| E Gene Clone a | ||
| E-1F | aaaggatccatgttcagctgtctggggatgcag | BamH I site (bold) |
| E-1R | aaggaattcggcattgacatttactgccaggaa | EcoR I site (bold) |
| BAC Insertion b | ||
| US2-F | aaagtcgacataacttcgtatagcatacattatacgaagttatcacgtcttcccgcgaggcc | Sal I site (bold), Lox p sequence (underline) |
| US2-R | ttaattaacgcggacaaaacgacgattac | Pac I site (bold) |
| SORF3-F | gtaatcgtcgttttgtccgcgttaattaatgaaaaagacggcggtacaat | Pac I site (bold) |
| SORF3-R | aagaatgcattcggcctgg | |
| Red-F | accagccagctgcgcttgctcgtggggtgtggtgcttttggt | |
| Red-R | tcgagcggccgctagggataacagggtaatccccaccttatatattctttcccaccctcgaagagcgttc | Not I site (bold), I-isce I sequence (underline) |
| Amp-F | aaattaattaaggggataacgcaggaaagaac | Pac I site (bold) |
| Amp-R | aaattaattaaacgtcaggtggcacttttcg | Pac I site (bold) |
| Modification pRThGA c | ||
| pCA-I-SceI-H1-F | aaatagggataacagggtaatgttgagcctttttgtggagtgggttaaattgtactagcgcgtttcgctttgcagtacatctacgtattagtcatcgctatta | I-isce I sequence (bold), Homology arm H1 (underline) |
| pCA-I-SceI-H2-R | aaatagggataacagggtaattagcatgcataacttcgtataatgtatgctatacgaagttatgcggccgc cacacaggaaacagctatgaccatgattac |
I-isce I sequence (bold), Homology arm H2 (underline) |
| Amp t-I-SceI-F | aaaattaccctgttatccctacacgttaagggattttggtcat | I-isce I sequence (bold) |
| OriT-R6K-I-SceI-R | aaaattaccctgttatcccta | I-isce I sequence (bold) |
| Identification E d | ||
| E-2F | atgttcagctgtctggggatgcag | |
| E-2R | ggcattgacatttactgccaggaa | |
a. Primers used to the clone the E gene of DTMUV; b. Primers used for the construction of the BAC insertion vector; c. Primers used for verification of the E gene inserted into the pBAC-C-KCE base using the MAGIC strategy; d. Primers used for the identification of the E gene of DTMUV.