TABLE 2.
Strains, plasmids, and primers used for the experiments in this study
| Strain, plasmid, or primer | Relevant features or sequencea | Reference(s) |
|---|---|---|
| Strains | ||
| Lactococcus lactis | ||
| KF147 | Plant-derived strain belonging to L. lactis subsp. lactis | 22 |
| NZ6200 | ΔcomEA-EC::tetR derivative of strain KF147 | This study |
| IL-9000 | Dairy-derived strain belonging to L. lactis subsp. lactis harboring nisRK integrated into its genome | 21, 45 |
| NZ9000 | Dairy-derived strain belonging to L. lactis subsp. cremoris harboring nisRK integrated into its genome | 29, 44 |
| KW2 | Plant-derived strain belonging to L. lactis subsp. cremoris | 28 |
| Escherichia coli TOP10 | Cloning host | |
| Plasmids | ||
| pIL253 | Emr; high-copy-number plasmid replicative in L. lactis | 43 |
| pNZ8040 | Cmr; pNZ123 derivative containing pepN downstream of the nisin promoter | 42 |
| pNZ8150 | Cmr; pNZ123 derivative with ScaI site downstream of the nisin promoter for translational fusion | 41 |
| pNZ6200 | Cmr; pNZ8150 derivative containing comX from L. lactis KF147 downstream of the nisin promoter | This study |
| pNZ6201 | Cmr; pNZ8150 derivative containing comX from L. lactis MG1363 downstream of the nisin promoter | This study |
| pNZ6202 | Tetr; pIL253 derivative with eryR replacement by tetR from pGhost8 | This study |
| pGhost8 | Tetr; vector with thermosensitive replicon, tetR | 69 |
| Primers | ||
| C1 | ATGACATATTACTTGGAAGAAGAGG | |
| C2 | CCTTGGTACCTCACTCTTCGTCTTCTGAAAATAAGATG | |
| C3 | ATGACATATTACCTGGAAGAAAATGAATTCG | |
| C4 | CCTTGGTACCTTAATCATCATCTCGAGAAAATAGTATATTTTTG | |
| C5 | AGATCTAGTCTTATAACTATACTGAC | |
| C6 | GCCTTGGTTTTCTAATTTTGGTTC | |
| C7 | GGAATGAAACGAGCAGATGCCC | |
| C8 | GTAATCATGGTCATAGCTGTTTCCACTTTTATATACGAAAAAACTCTTGGA | |
| C9 | TCCAAGAGTTTTTTCGTATATAAAAGTGGAAACAGCTATGACCATGATTAC | |
| C10 | GTCTTTTGCTCACTTTTTCCTTTCATGTTGTAAAACGACGGCCAGTG | |
| C11 | GGCACTGGCCGTCGTTTTACAACATGAAAGGAAAAAGTGAGCAAAAGAC | |
| C12 | ATTCATTGGAAGAAGACCTTTCGG | |
| C13 | CCCATAAAGCCGTAAACCAAGTGAAAG | |
| C14 | GAAGACCAAATTCTTTATTTTGCGG | |
| C15 | GATATCGAATTCCTGCAGCCCG | |
| C16 | CCTTAGTACTCTACAGAATATTACTATACACTCCAGAAG | |
| SS1 | CAGCGGAAGAGAACCGTATT | |
| SS2 | CTCAGTTCCTTGGATGCCAT | |
| PS1 | AGCAGCATAATAGATTTATTGAATAGG | |
| PS2 | GCATCTAATTTAACTTCAATTCCTATTATAC | |
| Q1 | CCTGGCGTACGTGAAGATGTC | |
| Q2 | TTTCGTCAGCCGGAACATAGC | |
| Q3 | TCTATTAGAAGAGCAGAGCGATGGTC | |
| Q4 | CTTGATAATGTGCGCTCAAGCCTTC | |
| Q5 | GTCAGCAGGCAAAGCTCTGTC | |
| Q6 | ACTTGACTAGTGACCGAATTAGCAGAG | |
| Q7 | ATGGCGACAACTATTTCCGAGCTCC | |
| Q8 | CAAGTTTGTCAGTAGAAGTTGCGGTC | |
| Q9 | CGCAGACGAGTTCAATTGGGAG | |
| Q10 | CGAGCCTACTGGATCAGCAAAGAG |
a
Underlining indicates restriction sites used in subsequent cloning procedures. Emr, erythromycin resistance; Cmr, chloramphenicol resistance; Tetr, tetracycline resistance.