Table 1

Bacterial strains and plasmids used in this study

Strain or plasmidRelevant characteristicsSource or reference
Strains
    LVSF. tularensis subsp. holarctica, type B, isolate (RG004) purified from a single colony, direct parent for genetic studiesT. Sellati, Albany Medical College, Albany, NY
    Schu S4F. tularensis subsp. tularensis, type A, isolate (CDC 1001) purified from a single colony, direct parent for genetic studiesCenters for Disease Control and Prevention, Ft. Collins, CO
    S4-029Schu S4 ΔiglE1::FRT-kan-FRTThis work
    S4-033Schu S4 ΔiglE1::FRTThis work
    S4-035Schu S4 ΔiglE1::FRT ΔiglE2::FRT-kan-FRTThis work
    S4-046Schu S4 ΔiglE1 ΔiglE2::FRTThis work
    S4-050Schu S4 ΔiglE1 ΔiglE2::FRT attTn7-iglEThis work
    S4-072Schu S4 ΔiglE1 ΔiglE2::FRT attTn7-iglE-Pro95 Δ(Asn95-Asp125)This work
    S4-074Schu S4 ΔiglE1 ΔiglE2::FRT attTn7-iglE (Cys22-Gly)This work
    TP367LVS ΔiglE1::FRT-kan-FRTThis work
    TP456LVS ΔiglE1::FRTThis work
    TP488LVS ΔiglE1::FRT ΔiglE2::FRT-kan-FRTThis work
    TP509LVS ΔiglE1 ΔiglE2::FRTThis work
    TP563TP509 with pTP181This work
    TP569LVS ΔiglE1 ΔiglE2::FRT attTn7-iglEThis work
    TP652LVS ΔiglE1 ΔiglE2::FRT attTn7-iglE-Pro95 Δ(Asn95-Asp125)This work
    TP656LVS ΔiglE1 ΔiglE2::FRT attTn7-iglE (Cys22-Gly)This work
    TP777LVS ΔiglE1 ΔiglE2::FRT attTn7-iglE Δ(Ser120-Asp125*a)This work
    TP809LVS ΔiglE1 ΔiglE2::FRT attTn7-iglE-Pro120 Δ(Ser120-Asp125)This work
    TP814LVS ΔiglE1 ΔiglE2::FRT attTn7-iglE-6× HisThis work
    TP816LVS ΔiglE1 ΔiglE2::FRT attTn7-iglE-Pro110 Δ(Ser110-Asp125)This work
    TP818LVS ΔiglE1 ΔiglE2::FRT attTn7-iglE-Pro126This work
    TP654LVS ΔiglE1 ΔiglE2::FRT attTn7-iglE-Pro2 Δ(Tyr2-Iso23)This work
Plasmids
    pMP590Suicide plasmid, Kanr SacB43
    pTP085Source of FRT-flanked PgroE-hyg-T1, HygrThis work
    pTP086Source of FRT-flanked PgroE-kan-T1, KanrThis work
    pTP101pMP590 with Kanr deleted and replaced with PgroE-hyg-T1 from TP085, HygrThis work
    pTP163pTP101 with oriT from pUC18T-mini-Tn7T, HygrThis work
    pTP181pMP829 with Tn7 helper functions (tnsABCD) from pTNS2, HygrThis work
    pLG66aSource of FRT-flanked Pfn-kan, Kanr42
    pTP363pTP163 + ΔFTT1701 (IglE)::FRT-Pfn-kan-FRT, Kanr HygrThis work
    pMP829Unstable shuttle vector, Hygr44
    pUC18T-mini-Tn7TTn7 delivery vector, oriT Ampr46
    pFFlp-hygTemperature-sensitive shuttle vector carrying Flp recombinase, Hygr42
    pTP405Unstable shuttle vector pMP829 expressing Flp under PrpsL control, HygrThis work
    pTP512pTP405 with sacB from pMP590, SacB HygrThis work
    pTNS2Source of Tn7 helper functions (tnsABCD), Ampr46
    pTP418pUC18T-mini-Tn7T with PrpsL-FTT1701 (iglE), AmprThis work
    pTP565pTP418 with FRT-flanked PgroE-kan-T1 from pTP086This work
    pTP795pTP418 iglE internal deletion control (adds Pro126), obtained by inverse PCR with GP301 and GP275This work
    pTP806pTP795 with FRT-flanked PgroE-kan-T1 from pTP086This work
    pTP631pTP418 with internal deletion of iglE C terminus, Δ(Asn96-Asp125) (adds Pro96), obtained by inverse PCR with GP274 and GP275This work
    pTP642pTP631 with FRT-flanked PgroE-kan-T1 from pTP086This work
    pTP792pTP418 with internal deletion of iglE C terminus, Δ(Ser110-Asp125) (adds Pro110), obtained by inverse PCR with GP300 and GP275This work
    pTP804pTP792 with FRT-flanked PgroE-kan-T1 from pTP086This work
    pTP789pTP418 with internal deletion of iglE C terminus, Δ(Ser120-Asp125) (adds Pro120), obtained by inverse PCR with GP299 and GP275This work
    pTP801pTP789 with FRT-flanked PgroE-kan-T1 from pTP086This work
    pTP750pUC18T-mini-Tn7T with PrpsL-iglE Δ(Ser120-Asp125*)This work
    pTP754pTP750 with FRT-flanked PgroE-kan-T1 from pTP086This work
    pTP744pUC18T-mini-Tn7T with PrpsL-iglE-6× HisThis work
    pTP808pTP744 with FRT-flanked PgroE-kan-T1 from pTP086This work
    pTP640pUC18T-mini-Tn7T + PrpsL-iglE (Cys22-Gly)This work
    pTP646pTP640 with FRT-flanked PgroE-kan-T1 from pTP086This work
    pTP635pTP418 with internal deletion of iglE signal sequence and putative lipobox, Δ(Tyr2-Iso23) (adds Pro2), obtained by inverse PCR with GP272 and GP273This work
    pTP644pTP635 with FRT-flanked PgroE-kan-T1 from pTP086This work
  • a The asterisk indicates that the construct lacks the nontemplated proline added by the inverse PCR mutagenesis strategy.