Quantification of plasmid loss in Escherichia coli cells by use of flow cytometry.
A method was developed to study plasmid stability in Escherichia coli cells, which utilised the high speed analysis properties of flow cytometry. To discriminate between plasmid-harbouring cells and plasmid-free cells a plasmid-encoded Lac repressor protein was used to regulate the expression of a chromosomally inserted green fluorescent protein gene in the host cells. Flow cytometric analysis enabled detection and quantification of plasmid-free cells due to their green fluorescent phenotype. The reported system offers real-time analysis in combination with a very low detection level of plasmid loss in bacterial populations. This could be useful in future investigations of plasmid stability and population selection in bacterial communities.
Bahl
Martin Iain
MI
Department of General Microbiology, University of Copenhagen, Sølvgade 83H, 1307 Copenhagen K, Denmark.
Sørensen
Søren Johannes
SJ
Hestbjerg Hansen
Lars
L
D015169:Colony Count, Microbial; D004926:Escherichia coli / Q000235:genetics* / Q000254:growth & development; D005434:Flow Cytometry; D015964:Gene Expression Regulation, Bacterial; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D008164:Luminescent Proteins / Q000235:genetics; D010957:Plasmids / Q000032:analysis*; D012097:Repressor Proteins
D008164:Luminescent Proteins; D012097:Repressor Proteins; D049452:Green Fluorescent Proteins