The hsp65 gene patterns of less common Mycobacterium and Nocardia spp. by polymerase chain reaction-restriction fragment length polymorphism analysis with capillary electrophoresis.
To rapidly identify Mycobacterium and Nocardia spp. without costly probes, we had implemented capillary electrophoresis (CE) in polymerase chain reaction (PCR)-restriction fragment length polymorphism (RFLP) analysis to analyze their 65-kDa heat shock protein (hsp65) gene. The PCR-RFLP analysis with CE (PRACE) involved only one restriction enzyme, HaeIII, and a single electrophoretic separation less than 10 min. Full-range (10-200 bp) RFLP patterns of 12 less common Mycobacterium and 7 Nocardia spp. were investigated. A good agreement was observed between the sizes of restriction fragments resolved by CE and the real sizes deduced from sequence analysis. Including hsp65 gene patterns of 12 Mycobacterium spp. published earlier, differentiation was distinct among 24 Mycobacterium and 7 Nocardia spp. Some closely related species exhibiting similar biochemical characteristics could be well discriminated by an extra HaeIII digestion site. Thus, PRACE offers a nonprobe alternative for rapid identification of various cultured Mycobacterium and Nocardia to the species level.
Chang
Po-Ling
PL
Department of Chemistry, National Taiwan University, Taipei, Taiwan.
Hsieh
Wen-Shyang
WS
Chiang
Chia-Lien
CL
Tuohy
Marion J
MJ
Hall
Gerri S
GS
Procop
Gary W
GW
Chang
Huan-Tsung
HT
Ho
Hsin-Tsung
HT
D001426:Bacterial Proteins / Q000235:genetics*; D018834:Chaperonin 60; D018833:Chaperonins / Q000235:genetics*; D004269:DNA, Bacterial / Q000235:genetics; D015252:Deoxyribonucleases, Type II Site-Specific; D019075:Electrophoresis, Capillary / Q000379:methods*; D009161:Mycobacterium / Q000145:classification* / Q000235:genetics; D009615:Nocardia / Q000145:classification* / Q000235:genetics; D016133:Polymerase Chain Reaction; D012150:Polymorphism, Restriction Fragment Length
D001426:Bacterial Proteins; D018834:Chaperonin 60; D004269:DNA, Bacterial; C074119:heat-shock protein 65, Mycobacterium; D015252:Deoxyribonucleases, Type II Site-Specific; C024403:GGCC-specific type II deoxyribonucleases; D018833:Chaperonins