Table 1.

Bacterial strains and plasmids used in this study

Strain or plasmidRelevant characteristicsSource or reference(s)
V. cholerae strains
 C6706Wild type, smooth, serogroup O1 El Tor biotype 34
 C6706 RC6706 rugose variantThis study
 569BWild type, smooth, serogroup O1 Classical biotype 12
 N16961Wild type, smooth, serogroup O1 El Tor biotype 28
 N16961 RN16961 rugose variantThis study
 NS1Derivative of N16961 R, smooth,epsD::TnphoA, KanR This study
 AA10N16961 R,epsD::Kanr cassetteThis study
 AA1N16961 R, epsE::KanrcassetteThis study
E. coli strains
 DH5α recA ΔlacU169 φ80dlacZΔM15Gibco BRL
 S17-1 λ pir pro hsdR hsdM + TmprStrr 7, 27
 SM10 λ pir thi thr leu tonA lacY supE recA::RP4-2Tc::Mu Kanr 7
Plasmids
 pBluescriptHigh-copy-number vector, Ampr,ori ColE1Stratagene
 pCVD442Suicide vector,ori R6K, Ampr,sacB 7
 pHC79Low-copy-number cosmid vector, Ampr, oriR6K 11
 pRT733TnphoA delivery vector, oriR6K, mob RP4, KanrAmpr 43
 pUC18 KHigh-copy-number vector, Ampr, ori ColE1, Kanr cassette mutagenesis vector 26
 pUC18 K2High-copy-number vector, Ampr,ori ColE1, Kanr cassette mutagenesis vectorJ. B. Kaper
 pAA5pHC79 containing a 38-kbSau3 AI fragment of NS1, AmprKanr This study
 pAA13pBluescript SK containing a 4.3-kb TnphoA insertion junction sequence from pAA5, Ampr This study
 pDSK-2pBluescript KS containing a 2.4-kb EcoRI-BamHI PCR fragment (epsD) from V. cholerae 569BThis study
 pAA26pBluescript SK containing a 2.4-kbBamHI-EcoRI PCR fragment (′epsD-5′-epsE′) from the N16961 rugose strainThis study
 pAA29pAA26::Kanr, a 0.8-kb Kanr cassette from pUC18K inserted into the uniqueNheI site at the 3′ end of epsD This study
 pAA35pCVD442 containing a 3.2-kbBamHI-EcoRI fragment from pAA29This study
 pAA40pBluescript SK containing 2.3-kbBamHI-KpnI PCR fragment (′epsD-epsE-epsF′)This study
 pAA42pAA40 containing Tetr cassette (EcoRI-AvaI fragment of pBR322) cloned in the XmnI siteThis study
 pAA46pAA42ΔepsE::Kanr, deletion of a HincII (bp 4117–4263) fragment and insertion of a 0.8-kb Kanr cassette from pUC18K2 into that siteThis study
 pAA48pCVD442 containing 3.1-kbBamHI-EcoRI fragment of pAA46 cloned into theSalI siteThis study