TABLE 1.

Strains, plasmids, and primers used in this study and their relevant characteristics

MaterialRelevant features or sequenceaSource or reference
Strains
    Escherichia coli MC1061Cloning host9
    Lactococcus lactis MG1363Cloning host18
    Lactobacillus plantarum
        WCFS1Wild-type for which the genome sequence is available28
        WCFS1-RRifrThis work
        NZ7109Rifr Emr; L. plantarum WCFS1 derivative containing a chromosomally located loxP-ery-loxP-cassetteThis work
Plasmids
    pUC18Apr54
    pUC19Apr54
    pNZ7105Apr; pUC18 derivative containing a 5′-truncated fragment of lp_3504This work
    pNZ7106Apr; pUC19 derivative containing a 5′-truncated fragment of lp_3503This work
    pNZ7107Apr; pNZ7105 derivative containing lp_3503 originating from pNZ7106This work
    pUC19lox1Apr; pUC18 derivative containing a loxP siteThis work
    pUC19lox2Apr; pUC18 derivative containing 2 loxP sites in tandem repeatThis work
    pGhost8Tetr35
    pNZ7103Apr Tetr; pUClox2 derivative containing tetR originating from pGhost8This work
    pNZ7108Apr Tetr; pNZ7107 derivative containing loxP-tetR-loxP cassette originating from pNZ7103This work
    pUC18eryApr Emr50
    pNZ7109Apr Emr; contains ery originating from pUC18ery flanked by two synthetic loxP sites and 5′-truncated fragments of lp_3503 and lp_3504 originating from pNZ7106 and pNZ7105, respectively (Fig. 1)This work
    pNZ7110Apr, pUC18 derivative containing cre and TpepN6
    pJIM2246Low-copy-cloning vector; Cmr44
    pNZ7124Cmr; pJIM2246 derivative containing cre and TpepN originating from pNZ7110This work
    pNZ7125Cmr; pNZ7124 derivative containing Tlox upstream of creThis work
    pNZ7126Cmr; pNZ7125 derivative containing PldhL1 upstream of creThis work
Primers
    cre-R25′-GTCCATCAGGTTCTTGCG-3′
    BglII-cre5′-ATAGTTTACCCCGTCAGC-3′
    lp_3503F5′-TGCTTTCCAAGAGCAAGCTG-3′
    lp_3503R5′-ACGTCTGCAGTCAGGTGTGAAGTTGGCACT-3′
    lp_3504F5′-ACGTCTGCAGCCACTCACCGACTAACACTC-3′
    lp_3504R5′-ACGTAAGCTTGACCCACGAGTTACCAACACG-5′
    flox15′-AGCTCTGCAGATAACTTCGTATAGCATACATTATACGAAGTTATTTGCA-3′
    flox25′-AATAACTTCGTATAATGTATGCTATACGTAGTTATCTGCAG-3′
    flox35′-CATAACTTCGTATAGCATACATTATACGAAGTTATCTGCAG-3′
    flox45′-AATTCTGCAGATAACTTCGTATAATGTATGCTATACGAAGTTATGAGCT-3′
    hrec35035′-GCATCGCTGCACTCTTTATG-3′
    hrec35045′-GAATATGGTCCATGGTCCAC-3′
    eryF5′-AAGCAATGAAACACGCC-3
    eryR5′-TTAGCCAGTTTCGTCGTT-3′
    lasF25′-ACGTGGATCCGGACAATATGGGGTAAGCG-3′
    lasR5′-AAGAAGATCTCTAAAGCTGACGGGGTAAAC-3′
    PldhL-F5′-GAAGATCTTCAATCTTCTCACCGTCTTG-3′
    PldhL-R5′-GAAGATCTTCAATAAGTCATCCTCTCGT-3′
  • a Underlined sequences indicate restriction sites subsequently used in cloning procedures. Apr, ampicillin resistant; Cmr, chloramphenicol resistant; Emr, erythromycin resistant; Rifr, rifampicin resistant; Tetr, tetracycline resistant.