Table 1.

Bacterial strains and plasmids

Bacterial strain or plasmidRelevant characteristicsSource or reference
E. coli
 DH5αλφ80d/lacZΔM15Δ(lacZYA-argF)U169 recA1 endA1 hsdR17(rK mK +)supE44 thi-1 gyrA relA1 GIBCO-BRL
 CJ236 dut ung thi relA; pCJ105 (Camr)Bio-Rad
 S17-1Tra genes from pRP4 integrated into chromosome for plasmid mobilization58
A. tumefaciens
 A136Strain C58 cured of pTi plasmid65
 A348A136 containing pTiA6NC31
 A348Spcr A348 with Spcr by spontaneous mutagenesisThis study
 PC1002A348 with virB2from pTiA6NC deleted7
 PC1004A348 with virB4from pTiA6NC deleted6
 PC1011A348 with virB11from pTiA6NC deleted7
Plasmid vectors
 pBSIISK+ NdeICrbr; cloning vector7
 pBSIIKS+ NdeICrbr; cloning vector7
 pSW172Tetr; broad-host-range IncP plasmid containing Plac with downstream polylinker sequence13
 pXZ151pSW172 with Kanr gene70
 pXZ1000KpSW172 ligated at KpnI site to ColE1 pBSKSK+ 69
 pML122ΔKmGenr; IncQ RSF1010 plasmid derivative with gentamycin resistance30
Expression plasmids
 pSR1Crbr; pBKS+ NdeI with virB11 expressed fromPvirB 51
 pJC902Crbr; pSR1 digested with SalI-XhoI and religatedThis study
 pJC903Crbr; pJC902 with XhoI site introduced after stop codon by site-directed mutagenesisThis study
 pJC8xxx seriespJC903 derivatives with 12-bp insertions corresponding to tandem NdeI-SalI sites immediately after codons indicated by xxxThis study
 pXZB100Crbr Kanr; pXZ151 withPvirB::virB11 69
 pXZB1xx seriesCrbr Kanr; pXZB100 with wild-typevirB11 gene replaced with PCR-mutagenized virB11alleles 1 through 1269
 pSRB9114CrbrTetr; pSW172 withPvirB::virB11 49
 pPCB7111Crbr Tetr; pSW172 withPlac::virB11 51
 pPCB7112Crbr Tetr; pSW172 withPlac::virB11ΔGKT174–176 51
 pPCB7113Crbr Tetr; pSW172 withPlac::virB11K175Q 51
 pZDH10Crbr Tetr; pSW172 withPlac::virB4 6
 pBBB15Crbr Tetr; pSW172 withPlac::virB4K439Q 6
 pBBB17Crbr Tetr; pSW172 withPlac::virB4ΔGKT438–440 6
 pVSB10Crbr Tetr; pSW172 withPvirB::virB2C64S 53