TABLE 6.
PCR primers applied during this study
| Primer nameb | Sequencea (5′–3′) | Target | Size of product (approx; bp) | Source |
|---|---|---|---|---|
| pNZ44F | CTAATGTCACTAACCTGCCCCG | pNZ44 MCS | Template dependent | This study |
| pNZ44R | GCTTTATCAACTGCTGCT | pNZ44 MCS | Template dependent | This study |
| STACas9iF | AGCAGCTCTAGAGTCGTTAGAGGGAGGATTAC | STA Cas9 | 3,366 | This study |
| STACas9iR | AGCAGCCTGCAGTTAAAAATCTAGCTTAGGC | STA Cas9 | This study | |
| STBCas9-1iF | AGCAGCTCTAGAGTTGCGAATTTTCAGATAC | STB CRISPR1-Cas9 | 3,366 | This study |
| STBCas9-1iR | AGCAGCCTGCAGGTAACTGTGTAAGGCGCC | STB CRISPR1-Cas9 | This study | |
| STBCas9-3iF | AGCAGCCTGCAGAAGGAGAAATGTATGACTAAG | STB CRISPR3-Cas9 | 4,167 | This study |
| STBCas9-3iR | AGCAGCCCATGGCTGGCTCTAGTTTAGGGTATT | STB CRISPR3-Cas9 | This study | |
| STBCsn2-1iF | AGCAGCCTGCAGCAGTGATAATAAGTTGGTGGT | STB CRISPR1-Csn2 | 1,053 | This study |
| STBCsn2-1iR | AGCAGCCCATGGCTGTCCTTGTCAATCCTTAC | STB CRISPR1-Csn2 | This study | |
| STBCsn2-3iF | AGCAGCTCTAGAGCCAATTCAGAGGAAAGG | STB CRISPR3-Csn2 | 660 | This study |
| STBCsn2-3iR | AGCAGCCTGCAGCAAGATGTGACTGTCACC | STB CRISPR3-Csn2 | This study | |
| CRISPR1F | TGCTGAGACAACCTAGTCTCTC | CRISPR1 repeat/spacer loci | Template dependent | 26 |
| CRISPR1R | TAAACAGAGCCTCCCTATCC | CRISPR1 repeat/spacer loci | Template dependent | 26 |
| CRISPR2F | TTAGCCCCTACCATAGTGCTG | CRISPR2 repeat/spacer loci | Template dependent | 26 |
| CRISPR2R | TAGTCTAACACTTTCTGGAAGC | CRISPR2 repeat/spacer loci | Template dependent | 26 |
| CRISPR3F | CTGAGATTAATAGTGCGATTACG | CRISPR3 repeat/spacer loci | Template dependent | 26 |
| CRISPR3R | GCTGGATATTCGTATAACATGTC | CRISPR3 repeat/spacer loci | Template dependent | 26 |
| CRISPR4F | GATTCAGTTCCTCATAGAGC | CRISPR4 repeat/spacer locus | Template dependent | This study |
| CRISPR4R | GACCTCAACCAATCGATTG | CRISPR4 repeat/spacer locus | Template dependent | This study |
| AC1g1 (internal) | CCTGTCATCTCTGGGAGT | STA CRISPR1 | NA | This study |
| AC1g2 (internal) | CGGTGTTCTATATCGAGGTC | STA CRISPR1 | NA | This study |
| AC1g3 (internal) | GTGAATGGGAAACTGACGGAA | STA CRISPR1 | NA | This study |
| AC3g1 (internal) | CAATCCGTAGCCACACCT | STA CRISPR3 | NA | This study |
| BC1g1 (internal) | CACTTGGCAGGCTTATTACTC | STB CRISPR1 | NA | This study |
| BC1g2 (internal) | CATCCGGTAACTGCTCAAGTG | STB CRISPR1 | NA | This study |
a
Incorporated restriction sites, where applicable, are underlined.
b
Internal gap-closing primers are indicated by a “g” and have been applied to all derivatives of the indicated parent S. thermophilus strain.