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Table 2 List of integration vectors and recombinant LAB strains

From: Development of a novel selection/counter-selection system for chromosomal gene integrations and deletions in lactic acid bacteria

Plasmid

Modified strain

Locus

Upstream region of homology

Downstream region of homology

Region deleted on chromosome/plasmid

Element(s) integrated onto chromosome/plasmid

pNZKOplaA::ErmFfluc

L. plantarum 423 plaA::erm-ffluc

plaA

902 bp upstream of plaA

69 bp internal fragment of plaA, 330 bp plaB bacteriocin immunity gene and 214 bp of plaC bacteriocin translocation protein

208 bp, including

102 bp of plaA

ORF

3204 bp erm-ffluc gene cassette

pNZKOmunA::CatFfluc

E. mundtii ST4SA munA::cat-ffluc

munA

911 bp upstream of munA, including 135 bp internal fragment of munA ORF

633 bp internal fragment of munB translocation gene

260 bp, including 18 bp of munA ORF

2471 bp cat-ffluc gene cassette

pNZKIaap::FRTerm

L. plantarum 423 aap::frt-erm

aap

909 bp internal fragment of aap ORF

929 bp internal fragment of aap ORF

None

1458 bp FRT-flanked erm gene

pNZKIaap::FRTerm

L. plantarum 423 aap::frt_um

 

909 bp internal fragment of aap ORF

929 bp internal fragment of aap ORF

1063 bp FRT-flanked erm resistance gene excised

335 bp including a 96 bp FRT-scar

pNZKOsrtA::FRTerm

E. mundtii ST4SA srtA::frt-erm

srtA

1011 bp upstream of srtA, including 57 bp internal fragment of srtA ORF, 333 bp mazG nucleotide pyrophosphohydrolase gene and 270 bp fragment of mfs transporter ORF

1011 bp downstream of srtA, including 57 bp internal fragment of srtA ORF and 576 bp internal fragment of bglG transcriptional anti-terminator ORF

580 bp internal fragment of the srtA ORF

1458 bp FRT-flanked erm gene

pNZKOsrtC::FRTerm

E. mundtii ST4SA srtC::frt-erm

srtC

1011 bp upstream of srtC, including 59 bp of srtC ORF and 892 bp of EbpCfm pilus subunit protein gene

376 bp downstream of srtA, including 60 bp of srtC ORF and 225 bp internal fragment of hemolysin III ORF

726 bp internal fragment of the srtC ORF

1458 bp FRT-flanked erm gene

  1. The integration vectors and their relevant characteristics are shown, and their construction is described in detail in the main text (pNZKOmunA::CatFfluc, pNZKIaap::FRTerm and pNZKOsrtA::FRTerm), Additional file 1: Text S1 (pNZKOplaA::ErmFfluc and pNZKOsrtC::FRTerm) and Additional file 1: Figs S2–S6. plaA: plantaricin 423 bacteriocin gene; erm: erythromycin resistance gene; ffluc; Photinus pyralis firefly luciferase gene; munA; mundticin ST bacteriocin gene; cat: chloramphenicol resistance gene; aap; FRT: flippase recombination target; L. plantarum 423 adhesion gene; um: unmarked; srtA: E. mundtii ST4SA sortaseA gene; srtC: E. mundtii ST4SA sortaseC gene