TABLE 1.

Bacterial strains and plasmids used in this study

Strain or plasmida	Genotype or characteristics	Source or reference
E. coli strains
BL21(DE3)	E. coli B, F−ompT hsdSB(rB− mB−) gal dcm (chromosomal T7 RNA polymerase on λDE3)	Novagen
BTH101	F−cya-99 araD139 galE15 galK16 rpsL1 (Strr) hsdR2 mcrA1 mcrB1	19
SPC105	MC4100 (ΔlacU169 araD rpsL relA thi flbB) containing a chromosomal PmarII::lacZ fusion at the λ attachment site and a wild-type mar locus	11
Plasmids
Two-hybrid system
pKT25	IPTG-inducible expression vector encoding the N-terminal T25 fragment (aa 1 to 224) of CyaA in frame with a multiple cloning site; Kmr; ori p15A	19
pUT18	IPTG-inducible expression vector encoding the C-terminal T18 fragment (aa 225 to 339) of CyaA in frame with multiple cloning site; Ampr; ori colE1	18
Two-hybrid analysis of MarR interactions
pUT18-marR	pUT18 derivative bearing the full-length marR as T18-ORF fusion	12
pKT25-gyrA	pKT25 derivative bearing the full-length gyrA as T18-ORF fusion	12
pKT25-marR	pKT25 derivative bearing the full-length marR as T25-ORF fusion	12
pKT25-marR R77H	pKT25-marR containing the Arg77His mutation of MarR	12
pTK25-marR G116S	pKT25-marR containing the Gly116Ser mutation of MarR	12
pTK25-marR SP34FS	pKT25-marR containing the Ser35Phe/Phe35Ser mutations of MarR	12
pTK25-marR D26N	pKT25-marR containing the Asp26Asn mutation of MarR	12
pTK25-marR G95S	pKT25-marR containing the Gly95Ser mutation of MarR	12
pTK25-marR L135F	pKT25-marR containing the Leu135Phe mutation of MarR	12
Analysis of tktA mutants
pSPOK	Kanamycin resistance derivative of pSPORT1; T7 promoter, lacO lacI	21
pSPOK-gyrA	pSPOK expressing gyrA	From Xiaowen R. Bina
pDrive	PCR cloning vector	Qiagen
pEGyr241/223	pUC18 with inducible external guide sequence 241/223 for GyrA	Gift from Sidney Altman (17)
pDrive-EGyr241/223	pDrive containing the inducible external guide sequences 241/223 for GyrA from pEGyr241/223	This study
His pulldown
pET13a	Expression cloning vector; Kanr; T7 promoter, lacO lacI	Novagen
pETmarR	marR cloned in pET13a	2
pET21b	Cloning vector for making C-terminal six-His fusion proteins; Ampr; T7 promoter, lacO lacI	Novagen
pETgyrA	pET21b containing gyrA	This study
Gyrase activity assay
pUC19 relaxed	Expression cloning vector; Ampr; relaxed by topoisomerase I of E. coli	New England Biolabs

^aThe strains used were E. coli K-12, unless noted otherwise in the text.