Compatibility of the green fluorescent proteinnext term and a general nucleic acid stain for quantitative description of a Pseudomonas putida biofilm

Nancharaiah, Y. V., Venugopalan, V. P., Wuertz, S., Wilderer, P. and Hausner, M. (2005) Compatibility of the green fluorescent proteinnext term and a general nucleic acid stain for quantitative description of a Pseudomonas putida biofilm. Journal of Microbiological Methods, 60 2: 179-187. doi:10.1016/j.mimet.2004.09.016


Author Nancharaiah, Y. V.
Venugopalan, V. P.
Wuertz, S.
Wilderer, P.
Hausner, M.
Title Compatibility of the green fluorescent proteinnext term and a general nucleic acid stain for quantitative description of a Pseudomonas putida biofilm
Journal name Journal of Microbiological Methods   Check publisher's open access policy
ISSN 0167-7012
Publication date 2005
Sub-type Article (original research)
DOI 10.1016/j.mimet.2004.09.016
Volume 60
Issue 2
Start page 179
End page 187
Total pages 9
Place of publication Amsterdam
Publisher Elsevier
Language eng
Subject 0605 Microbiology
Abstract Better understanding of biofilm development is essential for making optimal use of beneficial biofilms as well as for devising effective control strategies for detrimental biofilms. Analysis of biofilm structure and quantification of biofilm parameters using optical (including confocal) microscopy and digital image analysis techniques are becoming routine in many laboratories. The purpose of this study was to evaluate a dual labeling technique based on fluorescence signals from the green fluorescent protein (GFP) and those resulting from staining with the general nucleic acid stain SYTO 60 for the quantitative description of a model biofilm. For this purpose, a Pseudomonas putida KT2442 derivative was genetically tagged with the green fluorescent protein gene. Biofilm formation by this strain was investigated using flow cells and confocal laser scanning microscopy (CLSM). Percentage surface coverage as well as microcolony size quantified using GFP and SYTO 60 signals showed significant correlation (R=0.99). The results indicated that intrinsic labelling of this model biofilm using constitutively expressed proteins such as GFP can be used for real-time biofilm observation and generation of reliable quantitative data, comparable to those obtained using conventional methods such as nucleic acid staining. Non-destructive time series observation of GFP-expressing biofilms in flow-cells can thus be confidently used for four-dimensional (x, y, z, t) analysis and quantification of biofilm development. The results also point to the possibility of using GFP and SYTO 60 to study dual species biofilms, as quantitative data generated using both fluorophore signals are comparable.
Keyword Biofilm
CLSM
Dual fluorescence labeling
Green Fluorescent Protein
Pseudomonas putida
Q-Index Code C1
Q-Index Status Provisional Code
Institutional Status Non-UQ

Document type: Journal Article
Sub-type: Article (original research)
Collections: Excellence in Research Australia (ERA) - Collection
ERA 2012 Admin Only
Advanced Water Management Centre Publications
 
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