PMID:1100857

Citation	Aswad, D and Koshland, DE (1975) Isolation, characterization and complementation of Salmonella typhimurium chemotaxis mutants.J. Mol. Biol. 97:225-35
Abstract	No abstract in PubMed
Links	PubMed
Keywords	Chemotaxis; Genes, Dominant; Genes, Recessive; Genetic Complementation Test; Genotype; Mutation; Recombination, Genetic; Salmonella typhimurium; Diethylsulfate mutagenesis; Preformed liquid gradient technique
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Main Points of the Paper

Please summarize the main points of the paper.

• Isolation of non-chemotactic mutants
• Description of the preformed liquid gradient technique
  • utilizes a preformed gradient in a vertical column of liquid medium
  • separates wild-type, non-motile, and non-motile but chemotactic into 3 regions
• 72 mutants isolated
  • 68 exhibited smooth co-ordinated motility with no evidence of tumbling
  • 4 exhibited uncoordinated motility
• Results suggest there may be as many as 6 genetically distinct classes of che mutants in Salmonella

Materials and Methods Used

Please list the materials and methods used in this paper (strains, plasmids, antibodies, etc).

• For the isolation of non-chemotactic mutants
  • Diethylsulfate mutagenesis
  • Negative swarm selection on Tryptone semi-solid medium (0.3% agar)
  • Preformed liquid gradient technique (described in detail)

• For complementations and strain constructions
  • Alternative method of abortive transduction

• Strains
  • all strains were derived from Salmonella typhimurium LT2
  • Initial cultures ST2 and ST23 (wt chemotaxis) were both mutagenized with diethylsulfate to generate mutants

• For the initial ST2 culture

1. ST2 culture was mutagenized with diethylsulfate
2. culture spotted on plate
3. overnight incubation (wt bacteria swarmed outward from the inoculum, leaving a dense population at the center)
4. sample taken from the center of colony
5. perform 10 such serial selections
6. grow samples in nutrient broth and plate on Tryptone semisolid medium
7. 200 non-swarming colonies were picked and screened for motility by examination with a microscope

• For the ST23 culture

1. ST23 (wt chemotaxis) was mutagenized with diethylsulfate
2. che mutants enriched in the preformed liquid gradient apparatus 4 times, allowing for turbid growth between each enrichment
3. final culture plated on tryptone semisolid medium to screen for non-swarming colonies

3 distinct colony morphologies arose: (1)large colonies (wt); (2)tiny, compact colonies with sharp boundaries (non-motile); (3)tiny, compact colonies with fuzzy boundaries (che)

Phenotype Annotations

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Species	Taxon ID	Strain	Gene (if known)	OMP	Phenotype	Details	Evidence	Notes
Salmonella typhimurium LT2	NCBI:99287	ST20			Motility	consistently diffuses in the direction of gravity, regardless of attractant	Other	Preformed liquid gradient technique. Strain can also be labeled ST2che-222(p)
Salmonella typhimurium LT2	NCBI:99287	ST20			Motility	smooth motility with no spontaneous tumbling	Microscopy	isolated from using negative swarm selection described above. Phenotype labeled che-222(p)
Salmonella typhimurium LT2	NCBI:99287	ST1			Motility	moves in the direction of attractant	Other	Preformed liquid gradient technique
Salmonella typhimurium LT2	NCBI:99287	TA1859			Motility	no significant response to gradient	Other	Preformed liquid gradient technique. This is a non-motile mutant strain.
Salmonella typhimurium LT2	NCBI:99287	ST23che-223(e)			Motility	smooth motility in response to a temporal gradient of serine.	Microscopy	stroboscopic light source in dark field with an exposure time of 4s.
Salmonella typhimurium LT2	NCBI:99287	ST23che-223(e)			Motility	uncoordinated motility in either minimal medium of nutrient broth	Microscopy	stroboscopic light source in dark field with an exposure time of 4s.
Salmonella typhimurium LT2	NCBI:99287	ST23			Motility	wt chemotaxis
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Notes

• Controls for the first preformed liquid gradient experiment described in the paper were:

1. ST20
2. TA1859 (non-motile mutant)
3. ST1 (wild type strain)