Skip to main content
Journal of Bacteriology logoLink to Journal of Bacteriology
. 1981 Oct;148(1):391–393. doi: 10.1128/jb.148.1.391-393.1981

Enzymatic sorting of bacterial colonies on filter paper replicas: detection of labile activities.

C E Bulawa, B R Ganong, C P Sparrow, C R Raetz
PMCID: PMC216208  PMID: 6116698

Abstract

To utilize autoradiographic colony-sorting techniques (C. R. H. Raetz, Proc. Natl. Acad. Sci. U.S.A. 72:2274-2278, 1975) for the isolation of mutants with unstable enzymes, we report a new desiccation-induced lysis method, compatible with low temperatures. Furthermore, a general, two-step protocol is presented for clonal detection of hydrolytic reactions. The advantages of these critical modifications are demonstrated with the membrane enzymes glycerol 3-phosphate acyltransferase and cytidine 5'-diphosphate-diglyceride hydrolase.

Full text

PDF
391

Images in this article

Selected References

These references are in PubMed. This may not be the complete list of references from this article.

  1. Cramer C. L., Davis R. H. Screening for amino acid pool mutants of Neurospora and yeasts: replica-printing technique. J Bacteriol. 1979 Mar;137(3):1437–1438. doi: 10.1128/jb.137.3.1437-1438.1979. [DOI] [PMC free article] [PubMed] [Google Scholar]
  2. Esko J. D., Raetz C. R. Autoradiographic detection of animal cell membrane mutants altered in phosphatidylcholine synthesis. Proc Natl Acad Sci U S A. 1980 Sep;77(9):5192–5196. doi: 10.1073/pnas.77.9.5192. [DOI] [PMC free article] [PubMed] [Google Scholar]
  3. Ganong B. R., Leonard J. M., Raetz C. R. Phosphatidic acid accumulation in the membranes of Escherichia coli mutants defective in CDP-diglyceride synthetase. J Biol Chem. 1980 Feb 25;255(4):1623–1629. [PubMed] [Google Scholar]
  4. Langley K. E., Kennedy E. P. Partial purification and properties of CTP:phosphatidic acid cytidylyltransferase from membranes of Escherichia coli. J Bacteriol. 1978 Oct;136(1):85–95. doi: 10.1128/jb.136.1.85-95.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  5. Mowshowitz D. B. Permeabilization of yeast for enzyme assays: an extremely simple method for small samples. Anal Biochem. 1976 Jan;70(1):94–99. doi: 10.1016/s0003-2697(76)80051-6. [DOI] [PubMed] [Google Scholar]
  6. Mukherjee P. K., Paulus H. Biological function of gramicidin: studies on gramicidin-negative mutants. Proc Natl Acad Sci U S A. 1977 Feb;74(2):780–784. doi: 10.1073/pnas.74.2.780. [DOI] [PMC free article] [PubMed] [Google Scholar]
  7. Nishijima M., Raetz C. R. Membrane lipid biogenesis in Escherichia coli: identification of genetic loci for phosphatidylglycerophosphate synthetase and construction of mutants lacking phosphatidylglycerol. J Biol Chem. 1979 Aug 25;254(16):7837–7844. [PubMed] [Google Scholar]
  8. Raetz C. R., Dowhan W., Kennedy E. P. Partial purification and characterization of cytidine 5'-diphosphate-diglyceride hydrolase from membranes of Escherichia coli. J Bacteriol. 1976 Mar;125(3):855–863. doi: 10.1128/jb.125.3.855-863.1976. [DOI] [PMC free article] [PubMed] [Google Scholar]
  9. Raetz C. R. Enzymology, genetics, and regulation of membrane phospholipid synthesis in Escherichia coli. Microbiol Rev. 1978 Sep;42(3):614–659. doi: 10.1128/mr.42.3.614-659.1978. [DOI] [PMC free article] [PubMed] [Google Scholar]
  10. Raetz C. R. Isolation of Escherichia coli mutants defective in enzymes of membrane lipid synthesis. Proc Natl Acad Sci U S A. 1975 Jun;72(6):2274–2278. doi: 10.1073/pnas.72.6.2274. [DOI] [PMC free article] [PubMed] [Google Scholar]
  11. Raetz C. R. Isolation of Escherichia coli mutants defective in enzymes of membrane lipid synthesis. Proc Natl Acad Sci U S A. 1975 Jun;72(6):2274–2278. doi: 10.1073/pnas.72.6.2274. [DOI] [PMC free article] [PubMed] [Google Scholar]
  12. Raetz C. R., Newman K. F. Neutral lipid accumulation in the membranes of Escherichia coli mutants lacking diglyceride kinase. J Biol Chem. 1978 Jun 10;253(11):3882–3887. [PubMed] [Google Scholar]
  13. Raetz C. R. Phosphatidylserine synthetase mutants of Escherichia coli. Genetic mapping and membrane phospholipid composition. J Biol Chem. 1976 Jun 10;251(11):3242–3249. [PubMed] [Google Scholar]
  14. Snider M. D., Kennedy E. P. Partial purification of glycerophosphate acyltransferase from Escherichia coli. J Bacteriol. 1977 Jun;130(3):1072–1083. doi: 10.1128/jb.130.3.1072-1083.1977. [DOI] [PMC free article] [PubMed] [Google Scholar]

Articles from Journal of Bacteriology are provided here courtesy of American Society for Microbiology (ASM)

RESOURCES