An enhanced GFP reporter system to monitor gene expression in Borrelia burgdorferi.
Borrelia burgdorferi regulates genes in response to a number of environmental signals such as temperature and pH. A green fluorescent protein (GFP) reporter system using the ospC, ospA and flaB promoters from B. burgdorferi B31 was introduced into infectious clonal isolates of strains B31 and N40 to monitor and compare gene expression in response to pH and temperature in vitro. GFP could be assayed by epifluorescence microscopy, immunoblotting or spectrofluorometry and was an accurate reporter of target gene expression. It was determined that only 179 bp 5' of ospC was sufficient to regulate the reporter gfp in vitro in response to pH and temperature in B. burgdorferi B31. The loss of linear plasmid (lp) 25, lp28-1, lp36 and lp56 had no effect on the ability of B. burgdorferi B31 to regulate ospC in response to pH or temperature. The amount of OspC in N40 transformants was unaffected by changes in pH or temperature of the culture medium. This suggests that regulation of gene expression in response to pH and temperature may vary between these two B. burgdorferi strains.
Carroll
James A
JA
Rocky Mountain Microscopy Branch, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 903 South 4th Street, Hamilton, MT 59840, USA.
Stewart
Philip E
PE
Laboratory of Human Bacterial Pathogenesis, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 903 South 4th Street, Hamilton, MT 59840, USA.
Rosa
Patricia
P
Laboratory of Human Bacterial Pathogenesis, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 903 South 4th Street, Hamilton, MT 59840, USA.
Elias
Abdallah F
AF
Institut für Mikrobiologie und Hygiene, Charité Universitätsklinikum, Campus Charité Mitte Dorotheenstrasse 96, 10117 Berlin, Germany.
Garon
Claude F
CF
Rocky Mountain Microscopy Branch, Rocky Mountain Laboratories, National Institute of Allergy and Infectious Diseases, National Institutes of Health, 903 South 4th Street, Hamilton, MT 59840, USA.
D000942:Antigens, Bacterial; D000954:Antigens, Surface / Q000235:genetics; D019677:Artificial Gene Fusion; D001425:Bacterial Outer Membrane Proteins / Q000235:genetics; D001428:Bacterial Vaccines; D001483:Base Sequence; D025065:Borrelia burgdorferi / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D005408:Flagellin / Q000235:genetics; D015964:Gene Expression Regulation, Bacterial; D005798:Genes, Bacterial; D017930:Genes, Reporter; D049452:Green Fluorescent Proteins; D006863:Hydrogen-Ion Concentration; D008074:Lipoproteins; D008164:Luminescent Proteins / Q000235:genetics*; D008856:Microscopy, Fluorescence; D010957:Plasmids / Q000235:genetics; D011401:Promoter Regions, Genetic; D013045:Species Specificity; D013696:Temperature
D000942:Antigens, Bacterial; D000954:Antigens, Surface; D001425:Bacterial Outer Membrane Proteins; D001428:Bacterial Vaccines; D004269:DNA, Bacterial; D008074:Lipoproteins; D008164:Luminescent Proteins; C072045:OspA protein; C079616:OspC protein; D005408:Flagellin; C416532:flaB flagellin; D049452:Green Fluorescent Proteins