Flow Cytometric Testing of Green Fluorescent Protein-TaggedLactobacillus rhamnosusGG for Response to Defensins
| Autor: | Kristien Braeken, Mónica Perea Vélez, Tine Verhoeven, Sigrid C. J. De Keersmaecker, Sarah Lebeer, Jos Vanderleyden, Pascal Hols |
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| Rok vydání: | 2006 |
| Předmět: |
beta-Defensins
Green Fluorescent Proteins Applied Microbiology and Biotechnology Green fluorescent protein law.invention chemistry.chemical_compound Probiotic fluids and secretions Anti-Infective Agents Lactobacillus rhamnosus law In vivo Humans Gene Nisin Ecology biology Lacticaseibacillus rhamnosus Electroporation food and beverages Gene Expression Regulation Bacterial Flow Cytometry biology.organism_classification Molecular biology chemistry Food Microbiology Transformation Bacterial Food Science Biotechnology Transformation efficiency |
| Zdroj: | Applied and Environmental Microbiology. 72:4923-4930 |
| ISSN: | 1098-5336 0099-2240 |
| Popis: | Lactobacillus rhamnosusGG is of general interest as a probiotic. AlthoughL. rhamnosusGG is often used in clinical trials, there are few genetic tools to further determine its mode of action or to develop it as a vehicle for heterologous gene expression in therapy. Therefore, we developed a reproducible, efficient electroporation procedure forL. rhamnosusGG. The best transformation efficiency obtained was 104transformants per μg of DNA. We validated this protocol by taggingL. rhamnosusGG with green fluorescent protein (GFP) using the nisin-controlled expression (NICE) system. Parameters for overexpression were optimized, which allowed expression ofgfpinL. rhamnosusGG upon induction with nisin. The GFP+strain can be used to monitor the survival and behavior ofL. rhamnosusGG in vivo. Moreover, implementation of the NICE system as a gene expression switch inL. rhamnosusGG opens up possibilities for improving and expanding the performance of this strain. The GFP-labeled strain was used to demonstrate thatL. rhamnosusGG is sensitive to human beta-defensin-2 but not to human beta-defensin-1. |
| Databáze: | OpenAIRE |
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