[Detection of enterotoxigenic Escherichia coli by real-time polymerase chain reaction by using MGB-TaqMan probe technique].
OBJECTIVE
To develop a real-time polymerase chain reaction(PCR) based on TaqMan technology by using a new MGB probe for detecting enterotoxigenic Escherichia coli (ETEC) in paper.


METHODS
Primers and MGB probe were designed in the ecoding region of heat-stable toxin of ETEC. Real-time PCR detected ETEC by using the exterior standard method with protracting standard curves. The specificity, sensitivity, accuracy, stability of real-time PCR system was evaluated. An internal negative antithesis was added to the real-time PCR system in order to get rid of the false positive of system. Using UNG enzyme expelled the contamination of PCR reaction.


RESULTS
Primers and MGB probe were suited to the Real-time PCR. The assay showed that the method was quick, special, sensitive and stable. The real-time PCR system could detect ETEC in a large scale. The assay might be finished in two hour.


CONCLUSIONS
These observations suggested that real-time PCR based on MGB probe should be an excellent candidate for a standard ETEC detection method.
Dai
Juan
J
Bioengineering College, Xihua University, Chengdu 610039, China.
Li
Yu-feng
YF
Yuan
Li-xing
LX
Yang
Xiao
X
D001427:Bacterial Toxins / Q000302:isolation & purification*; D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial; D054307:Enterotoxigenic Escherichia coli / Q000302:isolation & purification*; D015336:Molecular Probe Techniques; D016133:Polymerase Chain Reaction / Q000379:methods*; D019914:Taq Polymerase
D001427:Bacterial Toxins; D017931:DNA Primers; D015342:DNA Probes; D004269:DNA, Bacterial; D019914:Taq Polymerase