Table 4 Bacterial strains^a and plasmids used in this study

E. coli S17-1 (λpir)

λpir (for replication of oriR6K-plasmids) recA, thi pro hsdR-M ⁺ RP4 2-Tc::Mu-Km::Tn7TpRSMR

[36]

P. fluorescens SBW25

Non-mucoid P. fluorescens wild type

[37]

SBW25MS1

Derivative of SBW25 where the Pm promoter is inserted directly upstream of algD using pMS9.

This study

SBW25MS1 ΔalgC:: TnKB61

Derivative of SBW25 MS1 where algC has been deleted utilizing pKB22, and a copy of algC controlled by the PmG5 promoter has been introduced on a transposon inserted into gene PFLU2944.

This study

SBW25mucAHE230

Alginate-producing derivative of SBW25 encoding a defect MucA and where the expression of algC is controlled by the PmG5 promoter

H. Ertesvåg, unpublished

SBW25mucAHE230ΔphoB

Derivative of SBW25mucAHE230 where an in-frame deletion in phoB was introduced utilizing pTK10.

This study

SBW25mucAHE230ΔphoR

Derivative of SBW25mucAHE230 where an in-frame deletion in phoR was introduced utilizing pTK9.

This study

Plasmids

pKD20

pUT based transposon vector containing PmG5. Ap^r, Km^r.

[5]

pLitmus28Tc

High copy number cloning vector. Tc^r, Ap^r

[5]

pMG48

RK2-based gene replacement vector. lacZ ⁺ , Tc^r, Ap^r

[33]

pMC1

RK2-based gene replacement vector for replacing the DNA sequence upstream of algD with the Pm-promoter. lacZ ⁺ , Tc^r, Ap^r

[10]

pKB22

Gene replacement vector for creating an algC-deletion. lacZ ⁺ , Tc^r, Ap^r

[5]

pKB60

Transposon vector. Contains the transposon TnKB60 with algC under the control of PmG5. Ap^r, Km^r

[5]

pYQ1

pUT based transposon vector containing PmG5. Am^r, Km^r.

[2]

pEM1

Derivative of pLitmus28Tc containing part of the transposon from pKD20. Ap^r

[2]

pKB61

Derivative of pKB60 where a 1.7 kb AvrII-NcoI DNA fragment encoding Km^r and most of XylS was exchanged with a 2.5 kb AvrII-NotI DNA fragment containing tetAR. Tc^r, Ap^r

This study

pMS9

Derivative of pMC1 where a 0.7 kb SbfI-NotI DNA fragment containing a gene upstream of algD was exchanged with a PCR product containing the 0.8 kb sequence directly upstream of algD. lacZ ⁺ , Tc^r, Ap^r

This study

pMS2

Derivative of pLitmus28Tc where the tetAR genes were exchanged with a 3.4 kb BamHI-fragment from pKD20 containing the minitransposon and oriR6K. Km^r, Ap^r

This study

pMS10

Derivative of pMS2 where a 0.5 kb BsiWI-EcoRI-fragment containing oriR6K was deleted and the 1.9 kb NotI-PstI fragment encoding XylS was exchanged with a 0.4 kb PCR product encoding oriR6K. Km^r, Ap^r

This study

pMS11

Derivative of pKD20 where a 3.7 kb BssHII-SfiI-fragment was exchanged with a 1.5 kb BssHII-SfiI-fragment containing oriR6K from pMS10. Km^r, Ap^r

This study

pTK1

Derivative of pEM1 in which a 2.2 kb PCR-amplified NdeI-NotI DNA fragment encoding phoBR from P. fluorescens was inserted. Ap^r.

This study

pTK3

Derivative of pTK1 in which an inserted 2.2 kb PCR-amplified NcoI-PspOMI DNA fragment from P. fluorescens including the first 46 nt of phoB replaced most of the phoB gene. Ap^r.

This study

pTK4

Derivative of pTK1 from which a 0.9 kb BstEII-BsaBI DNA fragment encoding most of phoR was deleted. Ap^r.

This study

pTK5

Derivative of pKD20 in which a 1.5 kb NdeI-NotI PCR fragment from P. fluorescens containing phoR was inserted. Km^r.

This study

pTK6

Derivative of pKD20 in which a 1.1 kb NcoI-NotI DNA fragment from pTK4 containing phoB was inserted. Km^r.

This study

pTK7

Derivative of pKD20 in which a 2.2 kb NdeI-NotI PCR fragment from P. fluorescens containing phoBR was inserted. Km^r.

This study

pTK8

Derivative of pTK1 from which a 0.3 kb BstEII DNA fragment was deleted, creating an in-frame deletion in phoR. Ap^r.

This study

pTK9

Derivative of pMG48 in which a 3.0 kb NcoI-NotI DNA fragment from pTK8 was inserted, containing a deletion in phoR. Ap^r, Tc^r.

This study

pTK10

Derivative of pMG48 in which a 3.1 kb NcoI-NotI DNA fragment from pTK3 was inserted, containing a deletion in phoB. Ap^r, Tc^r.

This study

pYQ1 trpF

Derivative of pYQ1 in which a 0.7 kb NdeI-NotI PCR fragment encoding TrpF was inserted. Am^r.

This study

pYQ1 trpD

Derivative of pYQ1 in which a 1.1 kb NdeI-NotI PCR fragment encoding TrpD was inserted. Am^r.

This study

pYQ1 trpDC

Derivative of pYQ1 in which a 1.9 kb NdeI-NotI PCR fragment encoding TrpDC was inserted. Am^r.

This study

pYQ1 purH

Derivative of pYQ1 in which a 1.6 kb NdeI-NotI PCR fragment encoding PurH was inserted. Am^r.

This study

pYQ1 purE

Derivative of pYQ1 in which a 0.5 kb NdeI-NotI PCR fragment encoding PurE was inserted. Am^r.

This study

pYQ1 ilvD

Derivative of pYQ1 in which a 2.1 kb NdeI-NotI PCR fragment encoding IlvD was inserted. Am^r.

This study

pYQ1 aceEI

Derivative of pYQ1 in which a 2.7 kb NdeI-NotI PCR fragment encoding AceE1 was inserted. Am^r.

This study

pYQ1 PFLU3030

Derivative of pYQ1 in which a 1.0 kb NdeI-NotI PCR fragment encoding PFLU3030 was inserted. Am^r.

This study

pYQ1 PFLU3887

Derivative of pYQ1 in which a 1.0 kb NdeI-NotI PCR fragment encoding PFLU3887 was inserted. Am^r.

This study

pYQ1 PFLU5567

Derivative of pYQ1 in which a 1.2 kb NdeI-NotI PCR fragment encoding PFLU5567 was inserted. Am^r.

This study

pYQ1 dsbC

Derivative of pYQ1 in which a 0.9 kb NdeI-NotI PCR fragment encoding DsbC was inserted. Am^r.

This study

pYQ1 sohB

Derivative of pYQ1 in which a 1.1 kb NdeI-NotI PCR fragment encoding SohB was inserted. Am^r.

This study

pYQ1 nagZ

Derivative of pYQ1 in which a 1.1 kb NdeI-NotI PCR fragment encoding NagZ was inserted. Am^r.

This study

pYQ1 anmK

Derivative of pYQ1 in which a 1.5 kb NdeI-NotI PCR fragment encoding AnmK was inserted. Am^r.

This study

pYQ1 ispA

Derivative of pYQ1 in which a 0.9 kb NdeI-NotI PCR fragment encoding IspA was inserted. Am^r.

This study

pYQ1 cbrB

Derivative of pYQ1 in which a 1.4 kb NdeI-NotI PCR fragment encoding CbrB was inserted. Am^r.

This study