TABLE 2.
Plasmids and strains used in this study
| Plasmid or strain | Descriptiona | Source or reference no. |
|---|---|---|
| Plasmids | ||
| pGEM-T Easy | TA cloning vector; Ampr | Promega |
| pProEX-HTb | Expression plasmid; N-terminal, TEV-cleavable His6 tag; Ampr | Invitrogen |
| pUAMS159 | pProEX-HTb::bosR; Ampr | This study |
| pUAMS338 | pProEX-HTb::cdaA (without the coding region for the putative transmembrane domains); Ampr | This study |
| pUAMS233 | pProEX-HTb::rpoS(120-131); Ampr | This study |
| pUAMS4 | pGEM-T Easy::PflgB-aacC1 (AscI flanked); Gentr, Ampr | 89 |
| pUAMS248B | ΔcdaA mutagenesis construct; Gentr, Ampr | This study |
| pUAMS309 | pGEM-T Easy::PflaB-aphI (BamHI flanked); Kanr, Ampr | This study |
| pUAMS313B | ΔcdaAC-cis complementation construct; Kanr, Ampr | This study |
| pUAMS351B | ΔcdaAC-cis(G175A) complementation construct; Kanr, Ampr | This study |
| pUAMS402 | pGEM-T Easy::PflaB-aadA (BamHI flanked); Spec/Strepr, Ampr | This study |
| pUAMS406B | ΔcdaAC-cis(G175A)::cdaAC-cis(WT) complementation construct; Spec/Strepr, Ampr | This study |
| pBSV2 | B. burgdorferi cp9-based shuttle vector; Kanr | 153 |
| pJD7 | Derivative of pKFSS1 B. burgdorferi shuttle vector; Spec/Strepr | 154 |
| pJD44 | pJD7-based shuttle vector with aph[3′]-IIIa; Kanr | 165 |
| pBtSV-JB | B. turicatae shuttle vector; Kanr | This study |
| pJSB104 | B. burgdorferi lac-inducible expression construct; Spec/Strepr | 154 |
| pUAMS267A | piRpoS, lac-inducible construct for BtWT rpoS; Kanr | This study |
| pUAMS346 | piCdaA, lac-inducible construct for BtWT bt0008; Kanr | This study |
| pUAMS446B | piBosR, lac-inducible construct for BtWT bosR; Kanr | This study |
| pRARE | Plasmid from Rosetta(DE3) cells; encodes rare tRNA codons in E. coli; Camr | Novagen |
| Strains | ||
| E. coli | ||
| TOP10F’ | F′ [lacIq Tn10 (Tetr)] mcrA Δ(mrr-hsdRMS-mcrBC) ϕ80lacZΔM15 nupG ΔlacX74 recA1 araΔ139 Δ(ara-leu)7697 galU galK rpsL (Strepr) endA1 | Life Technologies |
| C41(DE3) | F− ompT hsdSB (rB− mB−) gal dcm (DE3) | Lucigen |
| RosettaBlue(DE3) | endA1 hsdR17 (rK12− mK12+) supE44 thi-1 recA1 gyrA96 relA1 lac (DE3) F′[proA+ B+ lacIqZΔM15::Tn10] pRARE; Camr | Novagen |
| B. turicatae | ||
| BtWT | B. turicatae strain 91E135 (Oz1), tick isolate | 149, 150 |
| BtΔcdaA | cdaA null mutant, BtWT transformed with pUAMS248B; Gentr | This study |
| BtΔcdaAC-cis | cdaA null mutant complemented with a wild-type copy of cdaA (pUAMS313B); Gentr, Kanr | This study |
| BtΔcdaAC-cis(G175A) | cdaA null mutant complemented with cdaA containing the G175A point mutation (pUAMS351B); Gentr, Kanr | This study |
| BtΔcdaAC-cis(G175A)::cdaAC-cis(WT) | ΔcdaAC-cis(G175A) complemented with a wild-type cdaA allele (pUAMS406B); Gentr, Strepr | This study |
| BtiCdaA | BtWT transformed with pUAMS346; Kanr | This study |
| BtiBosR | BtWT transformed with pUAMS446B; Kanr | This study |
| BtiRpoS | BtWT transformed with pUAMS267A; Kanr | This study |
| BtiCdaA-ΔcdaA | Conditional B. turicatae cdaA mutant; BtiCdaA transformed with pUAMS248B; Gentr, Kanr | This study |
a
Amp, ampicillin; Gent, gentamicin; Kan, kanamycin; Spec, spectinomycin; Strep, streptomycin; Cam, chloramphenicol.