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Table 1 Plasmids, Strains and Oligonucleotides

From: Development of multiple strain competitive index assays for Listeria monocytogenes using pIMC; a new site-specific integrative vector

 

PLASMID

 

Name

Description

Reference

pTV1-OK

Temperature sensitive plasmid for the delivery of Tn917. 11 kb. EryR KanR.

[33]

pNZ272

Lactococcal beta-glucuronidase (gusA) transcriptional fusion vector. 4.6 kb. CmR.

[34]

pIMC

Site-specific Listerial integrative vector. High-level CmR from Phelp driven expression. 4.6 kb. CmR.

This study

pIMK3

Site-specific Listerial integrative vector. High level IPTG controlled gene expression. 7.5 kb. KanR.

[27]

pIMC3kan

Site-specific Listerial integrative vector. IPTG controlled expression of aphA-III ex pTV1-OK. CmR

[33] This study

pIMC3gus

Site-specific Listerial integrative vector. IPTG controlled expression of gusA ex pNZ272. CmR

[34] This study

pIMC3ery

Site-specific Listerial integrative vector. IPTG controlled expression of ermAM ex pTV1-OK. CmR

[33] This study

pIMC3tet

Site-specific Listerial integrative vector. IPTG controlled expression of tetM ex CH919. CmR

[35] This study

 

BACTERIA

 

Listeria monocytogenes

  

EGDe

Wild type 1/2a strain. Genome sequenced.

[9]

10403S

Wild type 1/2a strain. Genome sequenced. Broad Institute (MIT).

[36]

F2365

Wild type 4b strain. Genome sequenced.

[10]

EGDe::pIMC3kan

EGDe transformed with pIMC3kan integrated at the tRNAARG.

This study

EGDe::pIMC3gus

EGDe transformed with pIMC3gus integrated at the tRNAARG.

This study

EGDe::pIMC3ery

EGDe transformed with pIMC3ery integrated at the tRNAARG.

This study

EGDe::pIMC3tet

EGDe transformed with pIMC3tet integrated at the tRNAARG.

This study

10403S::pIMC3ery

10403S transformed with pIMC3ery integrated at the tRNAARG.

This study

F2365::pIMC3tet

F2365 transformed with pIMCtet integrated at the tRNAARG.

This study

Other Bacteria

  

DH10B

Wild type Escherichia coli. Used for routine cloning.

Invitrogen

CH919

MG1363 Lactococcus lactis subsp. lactis containing Tn919.

[36]

 

OLIGONUCLEOTIDE

 

Name

5'-3' a

RE site

108 Phelp B REV

CATGGGTTTCACTCTCCTTCTAC

 

181 CAT C FWD

AAATGTAGAAGGAGAGTGAAACCCATGAACTTTAATAAAATTGATTTAGAC

 

203 MCS FWD

AATTAACCCTCACTAAAGGGAAC

 

204 MCS REV

GACGTCGTAATACGACTCACTATAGGGC

 

205 Back FWD

AGTGAGTCGTATTACGACGTCCCAGGGCTTCCCGGTATCAAC

 

206 Phelp A FWD

ATCCCATTATGCTTTGGCAGTTTATTC

 

209 CAT D REV

ATAATGAGACAGAATTATGATGATCATATGTCAACTAACGGGGCAGG

 

211 Back REV

CATATGATCATCATAATTCTGTCTCATTATATAAC

 

373 gusA FWD

TATACCATGG TACGTCCTGTAGAAACCCCAACC

Nco I

374 gusA REV

ATATCTGCAG TTATTGTTTGCCTCCCTGCTGCGG

Pst I

385 aphA3 FWD

TAATCCATGG CTAAAATGAGAATATCACCGGAATTG

Nco I

386 aphA3 REV

AAGCCTGCAG TTAAAGCTTTTTAGACATCTAAATCTAGG

Pst I

387 ermAM FWD

ATATCCATGG ACAAAAATATAAAATATTCTCAAAAC

Nco I

388 ermAM REV

ATATCTGCAG TTATTTCCTCCCGTTAAATAATAGATAAC

Pst I

408 tetM FWD

ATATCCATGG AAATTATTAATATTGGAGTTTTAGC

Nco I

409 tetM REV

ATATCTGCAG TTAAGTTATTTTATTGAACATATATCG

Pst I

  1. a Restriction sites are in bold type