. Author manuscript; available in PMC: 2010 Mar 20.

Published in final edited form as: J Mol Biol. 2009 Jan 30;387(1):74–91. doi: 10.1016/j.jmb.2009.01.050

TABLE 4.

E. coli strains and plasmid DNAs used in this study.

E. coli strains
Strain	Relevant genotype	Source or construction
BL21(DE3)	F⁻ ompT hsdS _B(r_B⁻ m_B⁻) gal dcm (DE3)	Novagen
MS100	xyl-5 mtl-1 galK2 rpsL31 Δ(gpt-proA)62 lacY1 tsx-33 supE44 thi-1 hisG4(Oc) argE3(Oc) araD139 thr-1 sulA211 dnaN ⁺ tnaA300::Tn10	28
MS101	MS100: dnaN159 tnaA300::Tn10	28
AB1157	xyl-5 mtl-1 galK2 rpsL31 kdgK51 lacY1 tsx-33 supE44 thi-1 leuB6 hisG4(Oc) mgl-51 argE3(Oc) rfbD1 proA2 ara-14 thr-1 qsr-9 qin-111	CGSC^a⁴⁶
MS119	AB1157: dnaN⁺tnaA300::Tn10	19
MS120	AB1157: dnaN159 tnaA300::Tn10	19
MS134	AB1157: dnaN159 tnaA300::Tn10 ΔpolB::Ω	This work
MS123	AB1157: dnaN159 tnaA300::Tn10 Δ(dinB-yafN)::kan	19
MS122	AB1157: dnaN159 tnaA300::Tn10 ΔumuDC595::cat	19
MS135	AB1157: dnaN159 tnaA300::Tn10 ΔpolB::Ω Δ(dinB-yafN)::kan	This work
MS136	AB1157: dnaN159 tnaA300::Tn10 ΔpolB::Ω ΔumuDC595::cat	This work
MS124	AB1157: dnaN159 tnaA300::Tn10 Δ(dinB-yafN)::kan ΔumuDC595::cat	19
MS137	AB1157: dnaN159 tnaA300::Tn10 ΔpolB::Ω Δ(dinB-yafN)::kan ΔumuDC595::cat	This work
MS138	AB1157: dnaN⁺-tet-recF⁺	This work
MS139	AB1157: lamB::(His₆-dnaN⁺-cat)	This work
MS140	AB1157: ΔpolB::Ω Δ(dinB-yafN)::kan	This work
MS141	AB1157: ΔpolB::Ω Δ(dinB-yafN)::kan lamB::(His₆-dnaN⁺-cat)	This work
MS142	AB1157: ΔpolB::Ω Δ(dinB-yafN)::kan lamB::(His₆-dnaN⁺-cat) dnaN⁺-tet-recF⁺	This work
MS143	AB1157: ΔpolB::Ω Δ(dinB-yafN)::kan lamB::(His₆-dnaN⁺-cat) dnaN^148–152-tet-recF⁺	This work
MS144	AB1157: ΔpolB::Ω Δ(dinB-yafN)::kan dnaN⁺-tet-recF⁺ (pJD100)	This work
MS145	AB1157: ΔpolB::Ω Δ(dinB-yafN)::kan dnaN^148–152-tet-recF⁺ (pJD100)	This work
Plasmid DNAs
Plasmid	Relevant features	Source or construction
pET11a	Amp^R, ColE1 origin, directs overexpression of cloned gene under control of a T7 promoter	Novagen
pET11a-β5A	Amp^R, pET11a derivative overexpressing β^148–152 from the T7 promoter	This work
pβ^HMK	Amp^R, pET16b derivative overexpressing β⁺ with an N-terminal His₆ and kinase tag	47
pβ5A^HMK	Amp^R, pβ^HMK derivative overexpressing β^148–152	This work
pSU38	Kan^R, p15A origin, cloning vector	48
pACYCdnaN⁺	Kan^R, pSU38 derivative expressing physiological levels of β⁺	19
pACMβ5A	Kan^R, pSU38 derivative expressing physiological levels β^148–152	19
pACM	Cam^R, pSU38 derivative containing the cat gene inserted in kan	19
pACMdnaN⁺	Cam^R, pACM derivative expressing physiological levels of β⁺	19
pACMβ5A	Cam^R, pACM derivative expressing physiological levels of β^148–152	19
pAMP	Amp^R, pSU38 derivative containing the bla gene inserted in kan	19
pAMPdnaN⁺	Amp^R, pAMP derivative expressing physiological levels of β⁺	19
pAMPβ5A	Amp^R, pAMP derivative expressing physiological levels of β^148–152	19
pRD46	Amp^R, pSC101 origin, repA(Ts), expresses λ Red recombinase function under control of araBADC promoter	27
pAB1	Tet^R, pBR322 derivative containing 7.8-kbp PstIdnaA71 (Cs, Sx) dnaN⁺recF ⁺gyrB ⁺ cassette	49
pANTF	Kan^R Tet^R, pACYCdnaN⁺ derivative containing dnaA⁺-dnaN⁺-tet-recF⁺ cassette	This work
pANβ5TF	Kan^R Tet^R, pANTF derivative containing dnaA⁺-dnaN^148–152-tet-recF⁺ cassette	This work
pANβ5PVUIITF	Kan^R Tet^R, pANβ5TF derivative containing a copy of the dnaA⁺-dnaN^148–152-tet-recF⁺ cassette with a silent mutation within dnaN^148–152 that disrupts a PvuII restriction site	This work
pRMB100-Cat	Kan^R Cam^R, pSU38 derivative containing lamB::(His₆-dnaN⁺-cat) cassette	This work
pWSK29	Amp^R, pSC101 origin, low copy number cloning vector	50
pJD100	Amp^R, pSC101 origin, expresses physiological levels of dnaN⁺	28
pKOV	Cam^R, pSC101 origin, repA(Ts)	45
Oligonucleotides
Primer	Nucleotide sequence
80-mer	5′–[T₃₀]CTCCCTTCTTCTCCTCCCTCTCCCTTCCC[T₂₁]-Biotin–3′
30-mer	5′–AGGGAAGGGAGAGGGAGGAGAAGAAGGGAG–3′
SP20	5′–ACGCCTGTAGCATTCCACAG–3′
D150 loop top	5′–CTATGGCGGCTGCGGCCGCTGCCTATTACTAAATGG–3′
D150 loop bottom	5′–CCATTTAAGTAATAGGCAGCGGCCGCAGCCGCCATAG–3′
Tet top	5′–GCACCTCGAGTCAGCCCCATA–3′
Tet bottom	5′–GAGTGGTGAATTCGTTAGCG–3′
RecF top	5′–GAGCGCGAATTCTGTTGTCATGCC–3′
RecF bottom	5′–CAACGTTTCTCGAGCATTTATACTTGG–3′
D150ΔPvuII top	5′–CGTGCAACTAGAAGGTGAACGGATGCTGGTACGCTCCGG–3′
D150ΔPvuII bottom	5′–CCGGAGCGTACCAGCATCCGTTCACCTTCTAGTTGCACG–3′
JK28+2	5′–CATTGCCAATGCCAACTTTACC–3′
RecF back	5′–CTTCGAATTTTCGTCCGACATGTC–3′
DnaAP	5′–CATGAATGTTTCAGCCTTAGTC–3′
LamB-1	5′–CTCGCGCAAACTTCCTCTGGCGGTTGCCG–3′
LamB-2	5′–GAGACAAAGCTTGAGACGCGTGAGGTCGACGAGGGGTTGATTTCCATCTGCGCTAAACGCACATCG–3′
LamB-7	5′–GAGACAACGCGTGAGGGATCCACACGTGCCAACTTGCGTGATAACTATCGTCTGG–3′
LamB-8	5′–AGTGCCAAGCTTGAGCCATCTGGGCACCGAAGGTCCACTCGTC–3′
Cat promoter	5′–GAATTCGGATCCGTCTGCTATGTGGTGCTAT–3′
Cat end	5′–GAATTCGGATCCCTTATTCAGGCGTAGCACCAGGCG–3′
LamB-DN	5′–GAATGCGGCGTAAACGCCTTATCC–3′

CGSC, E. coli Genetic Stock Center, Yale University, New Haven CT, USA.