TABLE 1.

Bacterial strains and plasmids

Strain or plasmidCharacteristicsSource or reference
E. coli strains
    DH5αF φ80Δ lacZΔM15 Δ(lacZYA-argF)U169 endA1 recA1 hsdr17(rK mK+) deoR thi-1 supE44 gyrA96 relA1Life Technologies
    TOP10F mcrA Δ(mrr-hsdRMS-mcrBC) φ80lacZΔM15 ΔlacX74 nupG recA1 araD139 Δ(ara-leu)7697 galU galK rpsL(Strr) endA1 λInvitrogen
    BL21(DE3)(C43)F′ ompT hsdSB(rB mB) gal dcm (DE3)(C43)24
C. perfringens strains
    JIR32513 Nalr Rifr22
    EHE-NE18Australian chicken necrotic enteritis isolate20
    NE18ΔnetB1EHE-NE18 ΔnetB::catP19
    56Belgian chicken necrotic enteritis isolate14
    JIR4228JIR325 Ω Tn916pfoR pfoA colA luxS)5
    JIR12377EHE-NE18 virR Ω targetronThis study
    JIR1238256 virR Ω targetronThis study
Plasmids
    pMTL540FClostridial expression vector15
    pJIR750C. perfringens-E. coli shuttle vector, Cmr6
    pJIR750aipJIR750Ωalpha-toxin targetron8
    pJIR751C. perfringens-E. coli shuttle vector, Emr6
    pJIR1456C. perfringens-E. coli shuttle vector with oriT, Cmr21
    pJIR1877pUC18(EcoRI/HindII) harboring the product of PCR with UP/JRP372 (EcoRI/HindIII; 1.0 kb) containing virR and its promoter23
    pJIR1897pJIR750 Ω (HindIII/EcoRI: pJIR1877, 1.0 kb, virR)9
    pJIR2233pJIR750 (HindIII/BamHI) harboring the virR promoter (HindIII/BamHI: pJIR1877, 0.214 kb)This study
    pJIR2373pfoA reporter gene vector, Emr10
    pJIR2422pJIR2373(EcoRV) harboring annealed complementary primers 17443/17444 (34 bp) (wild-type pfoA VirR boxes)10
    pJIR2529pJIR2233(BamHI/SacI) harboring the virRS operon (BamHI/SacI; 2.1 kb)This study
    pJIR3464pJIR3754 (FspI/SpeI) harboring the replication region from pMTL540F (NheI/BstZ171; 1.73 kb)This study
    pJIR3562pJIR3756 (MluI) harboring PfdermB-RAM (MluI; 1.23 kb)This study
    pJIR3566Clostridial targetron vector derived from pJIR750ai, contains ermB-RAM and lacZα, CmrThis study
    pJIR3575pJIR2373(EcoRV) harboring annealed complementary primers JRP4051/JRP4052 (34 bp) (netB VirR boxes)This study
    pJIR3608Group II intron of pJIR3566 retargeted to the 546/547 site of the virR geneThis study
    pJIR3753pJIR750ai (HindIII/SphI) harboring Pgdh promoter region (HindIII/SphI; 207 bp)This study
    pJIR3754pJIR3753 (XbaI/SphI) harboring a PCR product containing the oriT of pJIR1456This study
    pJIR3756Clostridial vector harboring SOE PCR product (SalI/XbaI; 321 bp) containing the ferredoxin promoter (Pfd) fused to the ermB-RAM (PfdermB-RAM)This study