Table 1.

Strains and plasmids used in this study

Strain or plasmidGenotype and/or feature(s)Source or reference
Strains
 E2348/69Prototype O127:H6 EPEC strain 23
 JPN15E2348/69 cured of EAF plasmid 21
 UMD901E2348/69 bfpA, with S substituted for C at position 129 42
 CVD206E2348/69 Δeae 13
 UMD207E2348/69 Δeae, cured of the EAF plasmid 16
 UMD916E2348/69 bfpF::aphA3 This study
 DH5α supE44 ΔlacU169(φ80lacZΔM15) hsdR17 recA1 endA1 gyrA96 thi-1 relA1 30
 HB101 supE44 hsdSD20(rB mB )recA1 ara-14 proA2 lacY1 galK2 rpsL20 xyl-5 mtl-1 30
 DH5αλpir DH5α(λpir) 25
 BL21(DE3)F omp [lon]hsdSB (rB mB ; and E. coli B strain) with DE3, a λ prophage carrying the T7 RNA polymerase gene 35
Plasmids
 pBluescriptHigh-copy-number cloning vectorStratagene
 pCVD442Positive-selection suicide vector 13
 pDN19PBΔ1.3-kbPstI-BamHI fragment containing bfpPwith a 100-bp deletion cloned into pDN19 43
 pKDS135 bfp gene cluster cloned into pBR322 33
 pKDS134pKDS135 lackingbfpL 33
 pJPN14Fragment of EAF plasmid, required for expression of bfp genes, cloned into pACYC184 26
 pRPA53-kbSacI-EcoRI fragment containing bfpFcloned in pBluescriptThis study
 pRPA6pRPA5 with a 72-bpKpnI fragment deletedThis study
 pRPA102 bfpF cloned into low-copy-number vector pWKS30This study
 pUC18K aphA3 gene cloned into pUC18, creating a nonpolar kanamycin cassette 25
 pWKS30Low-copy-number cloning vector 39
 pRPAF1pKDS135 with 72-bp KpnI fragment deletedThis study
 pKDS8.1 Asp700 fragment containing bfpF::aphA3 cloned into pBluescriptThis study
 pKDS8.2 Asp700 fragment containing bfpF::aphA3 cloned into pCVD442This study
 pUMD916 bfpF::aphA3 EAF plasmid from UMD916This study
 pMSD205 bfpA gene cloned into pCR1000 under the control of an isopropyl-β-d-thiogalactopyranoside (IPTG)-inducible promoter 12