Abstract
The movement of bacteria through groundwater is a poorly understood process. Factors such as soil porosity and mineralogy, heterogeneity of soil particle size, and response of the bacteria to their environment contribute to the pattern of bacterial flow. The identification of transported bacteria is often a limiting factor in both laboratory and field transport experiments. Two bacterial strains were modified for use in bacterial transport experiments: a strain of Escherichia coli harboring the pGFP plasmid and a strain of Pseudomonas putida modified with a Tn5 derivative, Tn5GFP1. The Tn5GFP1 transposon incorporates the gene (gfp) encoding green fluorescent protein (GFP) and can be used to mutagenize Gram- bacteria. Fluorescent colonies were suspended in phosphate-buffered saline (PBS) at a concentration of approx. 109 bacteria/ml. A 10-cm glass column packed with quartz sand (diameter range 177-250 μm) was equilibrated with PBS prior to the forced flow introduction of the bacteria. Collected fractions were analyzed and the bacteria quantitated using a fluorescence spectrometer. Results demonstrate that the bacteria can be accurately tracked using their fluorescence, and that the intensity of the signal can be used to determine a C/Co ratio for the transported bacteria. The data show a rapid breakthrough of the bacteria followed by a characteristic curve pattern. A lower limit of detection of 105 cells was estimated based on these experiments. The Tn5GFP1 transposon should become a valuable tool for labeling bacteria.
Original language | English |
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Pages (from-to) | 53-58 |
Number of pages | 6 |
Journal | Gene |
Volume | 173 |
Issue number | 1 |
DOIs | |
State | Published - 1996 |
Funding
We thank L. Liang and J. McCarthyf or theire xcellent technicaal dvice.R esearchs upportedb y the Subsurface ScienceP rogramo f the Office of Health and Environmental Research,U S Departmenot f Energy, and by DefenseP rograms/USD epartmenot f Energy under a CooperativeR esearchand DevelopmentA greement (CRADA) No. DOE 92-0077b etweenL ockheedM arlin Energy ResearchC orp. and the National Center for ManufacturingS ciences.O ak Ridge National Labor-atoryis managefdo r the US Departmenotf Energyu nder contractD E-AC05-96OR2246w4 ith LockheedM arlin Energy ResearchC orp. PublicationN o. 4558,E nviron-mentalS ciencesD ivision,ORNL.
Keywords
- C/Co
- Soil column
- Tn5GFP1