Table 3 Bacterial strains and plasmids used in this work

Rhizobium leguminosarum

UPM791

128C53 wild type; Str^r Nod⁺ Fix⁺ Hup⁺

[40]

UPM1155

UPM791 ( Δhup/hyp cluster) Hup^-

[19]

Escherichia coli

DH5_α

recA1 endA1 gyrA96 thi hsdR17 supE44 relA1 Δ(lacZYA-argF)U169 Φ80dlacZΔM15

[41]

S17.1

thi pro hsdR^-hsdM⁺recA RP4::2-Tc::Mu-Kan::T7; (Sp^r Sm^r)

[42]

Plasmids

pAL618

pLAFR1-based cosmid containing the whole R. leguminosarum hydrogenase gene cluster

[40]

pALPF1

pAL618 with hupSL promoter replaced by fixN promoter (P _fixN )

[18]

pALPF2

pALPF1 ΔhupL

[19]

pALPF4

pALPF1 ΔhupD

[19]

pALPF5

pALPF1 ΔhupF

This work

pALPF10

pALPF1 ΔhupK

This work

pALPF14

pALPF1 ΔhypC

This work

pALPF382

pALPF1 derivative carrying hupF _ST gene

This work

pBBR1MCS-2

Broad-host-range plasmid; Km^r mob⁺

[43]

pKD3

Template plasmid harbouring FLP-mediated excision sequences flanking Cm^r gene

[44]

pPM71

PKD3 derivative containing Strep-tag II sequence for C-terminal end fusion

This work

pPM1350

pBBR1MCS-2 derivative containing a DNA fragment harbouring P _fixN promoter from R. leguminosarum

[19]

pPM501

pPM1350 derivative containing an NdeI-XbaI fragment harbouring HupF _ST under the control of P_fixN

This work

pPM501C

pPM501 derivative containing a deletion of the 25 3′codons of hupF

This work

pPCR2.1-TOPO

PCR cloning vector

Invitrogen