Open AccessSimplified procedures for detection of amplified DNA using fluorescent label incorporation and reverse probing
Author(s) -
Woolford Alison J.,
Dale Jeremy W.
Publication year - 1992
Publication title -
fems microbiology letters
Language(s) - English
Resource type - Journals
SCImago Journal Rank - 0.899
H-Index - 151
eISSN - 1574-6968
pISSN - 0378-1097
DOI - 10.1111/j.1574-6968.1992.tb05587.x
Subject(s) - polymerase chain reaction , fluorescence , microbiology and biotechnology , dna , primer dimer , hybridization probe , fluorescein , chemistry , multiple displacement amplification , real time polymerase chain reaction , biology , computational biology , multiplex polymerase chain reaction , biochemistry , dna extraction , gene , physics , optics
Conventional methods of detecting polymerase chain reaction (PCR) products require equipment and expertise which may not be available in diagnostic bacteriology laboratories, especially in developing countries. To this end we have examined other methods of product detection, including fluorescein‐12‐dUTP incorporation during PCR amplification, and reverse probing, where the PCR product is used as the probe in a scaled down hybridization with a fixed capture probe consisting of a fragment entirely internal to the sequence of the PCR product. These techniques have shown sensitivities of 20 fg of purified mycobacterial DNA, which corresponds to approximately five cells.