Table 2.

Comparison of reagents and conditions between theE. coli O157:H7 PFGE protocol and the rapid PFGE protocol for Campylobacter

Reagent or conditionsReagent and/or condition(s) for protocola:
PulseNet protocol for E. coli O157:H7Campylobacter rapid PFGE protocol
CSBTE: 100 mM Tris, 100 mM EDTA, pH 8.0PBS: 0.01 M phosphate buffer (pH 7.4), 0.85% NaCl
Plug agarose1% SKG–1% SDS in TE1% SKG in TE
Lysis buffer50 mM Tris, 50 mM EDTA (pH 8.0), 1% sarcosine, 0.1 mg of proteinase K/ml50 mM Tris, 50 mM EDTA (pH 8.0), 1% sarcosine, 0.1 mg of proteinase K/ml
Washes (10 to 15 min each)Six at 50°C (twice with sterile ultrapure water and four times with TE)Four at 54°C (once with sterile ultrapure water and three times with TE)
Restriction digestion25 to 50 U (XbaI) at 37°C for 2 h40 U (SmaI) at 25°C for 2 h
Electrophoresis conditionsIST = 2.16 s; FST = 54.17 s; gradient = 6 V/cm; included angle = 120; running time = 18 to 19 hIST = 6.75 s; FST = 38.35; gradient = 6 V/cm; included angle = 120; running time = 18 to 19 h
  • a IST, initial switch time; FST, final switch time.