Table 4 Bacterial strains and plasmids used in this study

Plasmid

pJET1.2

Ap^r, rep (pMB1) from pMBI responsible for replication

Thermo scientific

pCAM-MCS

Ap^r, pCAM140-derivative without mini-Tn5, contains the MCS of pBluescript II SK (+)

[16]

pFCm1

Ap^r, Cm^r, source of Cm^r cassette flanked by FRT sequences

[43]

pCAM-Km

Km^r, variant of the gene replacement vector pCAM-MCS, Ap^r replaced by Km^r

This study

pCAM-Km.acrD-Cm

Km^r, Cm^r, contains a 1.1-kb fragment of acrD from E. amylovora Ea1189, insertion of 1153-bp Cm-FRT cassette from pFCm1 in BamHI site

This study

pBlueScript II SK(+)

Ap^r, ColE1 origin

Stratagene

pBlueScript II KS(+)

Ap^r, ColE1 origin

Stratagene

pBBR1MCS

Cm^r, ColE1 origin

[44]

pBlueKS.acrD

Ap^r, contains a 3.1-kb fragment carrying acrD of E. amylovora Ea1189 under control of lac promoter

This study

pBlueKS.acrD-ext

Ap^r, contains a 3.5-kb fragment carrying acrD of E. amylovora Ea1189 including promoter region under control of lac promoter

This study

pBlueSK.acrD

Ap^r, contains a 3.1-kb fragment carrying acrD of E. amylovora Ea1189 in opposite orientation with respect to lac promoter

This study

pBlueSK.acrD-ext

Ap^r, contains a 3.5-kb fragment carrying acrD of E. amylovora Ea1189 including promoter region in opposite orientation with respect to lac promoter

This study

pBBR.egfp.TIR

Cm^r, contains the TIR-egfp-T₀ cassette in pBBR1MCS in opposite orientation with respect to lac promoter

[16]

pBBR.acrD-Pro.egfp

Cm^r, contains a 206-bp fragment carrying the promoter region of acrD, transcriptional fusion of acrD with egfp

This study

pBBR.acrA-Pro.egfp

Cm^r, contains a 133-bp fragment carrying the promoter region of acrA, transcriptional fusion of acrA with egfp

This study

pBlueSK.baeR

Ap^r, contains a 0.7-kb fragment carrying baeR of E. amylovora Ea1189 under control of lac promoter

This study

pET-28a(+)

Km^r, f1 origin

Novagen

pET28a.baeR

Km^r, contains a 0.7-kb fragment carrying baeR of E. amylovora Ea1189, C-terminal translational fusion with His-tag

This study

pCP20

Cm^r, Ap^r, contains yeast Flp recombinase gene, rep (pSC101) responsible for temperature-sensitive replication

[45]

pBAD24

Ap^r, pMB1 origin, araC

[46]

pBAD24.baeR

Ap^r, contains a 0.7-kb fragment carrying baeR of E. amylovora Ea1189 under control of P_BAD promoter

This study

Strain

Escherichia coli

XL1-Blue

endA1 gyrA96(nal^R) thi-1 recA1 relA1 lac glnV44 F'[ ::Tn10 proAB⁺ lacI^q Δ(lacZ)M15] hsdR17(r_K ^- m_K ⁺)

Stratagene

TG1

K-12 supE thi-1 Δ(lac-proAB) Δ(mcrB-hsdSM)5, (r_K ^-m_K ^-)

[47]

KAM3

acrAB mutant of TG1

[48]

BL21(DE3)

F^– ompT gal dcm lon hsdS_B(r_B ^- m_B ^-) (λDE3)

Novagen

S17-1

TpR SmR recA, thi, pro, hsdR-M⁺RP4: 2-Tc:Mu: Km

[49]

S17-1 λ-pir

λpir phage lysogen of S17-1

[49]

DH5α λ-pir

sup E44, ΔlacU169 (ΦlacZΔM15), recA1, endA1, hsdR17, thi-1, gyrA96, relA1, λpir phage lysogen

D. Lies, Caltech

Erwinia amylovora

Ea1189

Wild type

GSPB^b

Ea1189-3

Km^r, acrB mutant carrying Km^r cassette in the acrB gene

[16]

Ea1189.acrD

acrD mutant

This study