An improved protocol for the preparation and restriction enzyme digestion of pulsed-field gel electrophoresis agarose plugs for the analysis of Legionella isolates.
Pulsed-field gel electrophoresis (PFGE), which determines the genomic relatedness of isolates, is currently used for the epidemiological investigation of infectious agents such as bacteria. In particular, this method has been used for the epidemiological investigation of Legionella outbreaks. However, it takes 4 days to complete a Legionella-PFGE analysis. Due to partial digestion and DNA damage, the reproducibility of the obtained fragment digestion patterns is poor for this pathogen. In this study, we report an improved protocol that takes only 2 days to complete and that allows clear discrimination of the restriction profile with higher reproducibility than that previously achieved.
Chang
Bin
B
Department of Bacteriology, National Institute of Infectious Diseases, Tokyo 162-8640, Japan.
Amemura-Maekawa
Junko
J
Watanabe
Haruo
H
D000818:Animals; D015373:Bacterial Typing Techniques; D004262:DNA Restriction Enzymes / Q000032:analysis*; D004196:Disease Outbreaks; D016521:Electrophoresis, Gel, Pulsed-Field / Q000379:methods*; D007875:Legionella / Q000145:classification* / Q000235:genetics / Q000302:isolation & purification; D015203:Reproducibility of Results; D012685:Sepharose / Q000032:analysis; D012703:Serotyping
D012685:Sepharose; D004262:DNA Restriction Enzymes