KIT 4

F Prime Factors with DNA of E. coli in S. typhimurium

This kit includes strains of S. typhimurium LT2 which carry F-prime factors which carry genes derived from E. coli K-12, representing a large part (but not all) of the chromosome of E. coli. They were constructed originally by Brooks Low, and transferred into S. typhimurium by conjugation. These conjugates were originally reported by Sanderson and Hartman,1978, Microbiol. Revs 42:417-519. Later L. Csonka and colleagues inserted transposons bearing antibiotic resistance to the F-prime factor, in order to increase the value of these F-prime factors for genetic analysis.

The strains are described in Sanderson and Roth, 1988, Microbiol. Revs 52:485-532. The chromosome of these strains is S. typhimurium LT2; all are recA+, but the fact that the DNA on the F-prime factor is from E. coli and has low homology with S. typhimurium DNA means that the probability of insertion of the F-prime factor is low.

In about half of the18 strains the F-prime factor is wild type for replication; in the other half it is temperature-sensitive, i.e. unable to replicate normally at temperatures significantly higher than 30 degrees C. These strains must be grown at 30 degrees C in broth, otherwise the F-prime factor will be lost unless selective pressure is applied, and even then strains in which the F-prime factor is inserted into the chromosome will likely be isolated. These strains can be used with selection at high temperature to obtain Hfr strains.

The F-prime factor can be retained in the strain by growing in the presence of the antibiotic (tetracycline for Tn10, kanamycin for Tn5). Transfer of the antibiotic resistance by conjugation can be selected directly, and other genes on the F-prime factor can be detected as unselected markers.

List of strains of S. typhimurium with F-prime factors which carry chromosomal genes of E. coli K-12, as well as transposon Tn5 or Tn10.

Additonal genetic information can be found by searching the SGSC on-line search site (Start new search)

 Strain #

SGSC#

F' factor

Approx posn. of factor on E. coli map (min) (a)

 Genes confirmed on F' (b)

 Chromosomal markers

Phenotype of strain

Mutations complemented in this strain by F' genes

Temperature sensitive
 TL851 1370 F104 97-7  leu+ proB+ A+ zzf-20::Tn10  purB655 (del)proBA47 PyrB- (Ura) Tc R Pro+ R
 TL852 1371 F'104  97-7  leu+ proB+ A+ zzf-20::Tn10  purB655 (del)proBA47 PyrB- (Ura) Tc R  Pro+  S
 TL873 1372  F'128  6-8 ProB+ A+ zzf-20::Tn10 argF+ lacI4000phi(lacI-Z) Y+ argI537 ara-9 fol-1 Ara- Fol- Tc R  Arg+ Lac +  R
 TL874 1373  F'128  6-8 ProB+ A+ zzf-20::Tn10 argF+ lacI4000phi(lacI-Z) Y+  argI537 ara-9 fol-1 Ara- Fol- Tc R  Arg+ Lac +  S
 TL275 1374   F'128 6-8 ProB+ A+ argF+ lacI4000phi(lacI-Z) Y+, Tn5 in unknown location on F'128  (del)proBA21 pyrA8 argR5 fol-181 rpsL201 PyrA- (arg+ Ura) Fol- Sm R Km R  Pro+  R
 TT1948   323  F'152 12-17 nadA+ zzf-20::Tn10 his0124 hisB2142 nad-506 hut+ galE542 His- Tc R  Nad+ Gal+  R
 TL853  1375  F'148  32-34, 42-44  his+ zzf-20::Tn10 trpA49 pncA15 hisD9953::MudI1734 Trp- PncA- Km R Tc R  His+  R
 TL854  1376  F'148  32-34, 42-44  his+ zzf-20::Tn10 trpA49 pncA15 hisD9953::MudI1734 zcc-628::Tn5 Trp- PncA- Km R Tc R  His+  S
 TL1600  1533  F'129  44-51  his+ zzf-20::Tn10 his-2236 proC1909::Mud1-8 rpsL1 His+ Pro- Sm R His+  S
 TL1178  1390  F'198  50-56 cysA+ zzf-20::Tn10 thr-469::Mud1-8 Thr- Ap R Tc R  -  R
 TL1179  1391  F'198  50-56 cysA+ zzf-20::Tn10 thr-469::Mud1-8 Thr- Ap R Tc R  -  S
 TL870  1392  F'143  56-62 cysC+ zzf-20::Tn10 cysC519 rpsL1 Sm R Tc R Cys+   R
 TL871  1393  F'143  56-62 cysC+ zzf-20::Tn10 cysC519 rpsL1 Sm R Tc R  Cys+   S
 TL863  1394  F'116  59-66 lysA+ serA+ zzf-20::Tn10 lys554 serA790 his-644 His- Tc R Lys+ Ser+  R
 TL864  1395  F'116  59-66 lysA+ serA+ zzf-20::Tn10 lys554 serA790 his-644 His- Tc R  Lys+ Ser+  S
 TL860  1396  F'140  67-81 cysG+ argD+ zzf-20::Tn10 argD455 (del)proBA rpsL1 Pro- Sm R Tc R  Arg+  S
 TL865  1397  F'117  94-97 purB+ zzf-20::Tn10 pyrB64 hisD6414 mel His- Tc R PyrB+  R
 TL866  1398  F'117  94-97 purB+ zzf-20::Tn10 pyrB64 hisD6414 mel His- Tc R  PyrB+  S

(a) The F-prime factors of E. coli wre originally described by Low (1972. Bacteriol. Rev. 36: 587-607). However, the location of the genes carried is given according to edition VII of the E. coli Map (Bachmann, B.J., 1983. Microbiol. Rev. 47:180-230) in which the minutes have been modified

(b) The temperature-sensitive mutation in F' ts114 lac (Jacob et al., 1963. Cold Springs Harbour Symp. Quant. Biol. 28:329-348, which makes the plasmid unable to replicate at high tempeature, is linked by P22 transduction to the transposon insertion zzf-20::Tn10 (constructed by Chumley et al., 1979. Genetics 91: 639-655). The new F factors were made by transducing from an F-prime factor carrying both ts114 and zzf-20::Tn10 into the E. coli F factors. All isolates selected carried the transposon; some are temperature sensitive (carry the ts allele), whereas others are temperature resistant. Those which are temperature sensitive must be grown at 30 degrees C to prevent loss of the plasmid or insertion of the plasmid into the chrommosome.

References:

Chumley, F.G., Menzel, R., and Roth, J.R. 1979. Hfr formation directed by Tn10. Genetics 91:639-655.

Sanderson, K.E. and Hartman, P.E. 1978. Linkage map of S. typhimurium, edition V. Microbiol. Rev. 42:471-519

Sanderson, K.E. and Roth, J.R. 1988. Linkage map of S. typhimurium, edition VII. Microbiol. Rev. 52:485-532.

Sanderson, K.E. 1996. F-mediated conjugation, F+ strains, and Hfr strains of Salmonella typhimurium and Salmonella albony. In E. coli and Salmonella, editon 2, F.C. Neidhardt et at., (editors), ASM press, pps 2406-2412

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