New molecular probe for real-time PCR monitoring and genetic testing
13 August 2013
Eprobe is a fluorescent probe for PCR DNA amplification
techniques and DNA analysis developed by researchers from RIKEN and
Japanese firm K.K.DNAForm.
The probe will enable the development of new, advanced assays for
DNA-based genetic testing and help to bring the benefits of
genome-wide sequencing studies to patients in the clinic.
PCR, for Polymerase Chain Reaction, is a simple and inexpensive
DNA amplification technique, widely used to analyse DNA and RNA in
life science laboratories. PCR is also used in hospitals to diagnose
diseases, identify bacteria and viruses, or in forensic medicine.
During PCR, small amounts of target DNA molecules are copied and
rapidly amplified, thus enabling researchers to analyze the DNA,
test it or clone it.
Eprobes are short DNA oligonucleotides labelled with two
fluorescent dye moieties attached to the same nucleotide. During the
DNA amplification reaction, the probes bind to the newly synthesized
DNA fragments, emitting a strong fluorescence signal upon binding,
which enables researchers to monitor the reaction in real-time.
In the study, the researchers used Eprobes for the detection of
genetic variations in the human EGFR and KRAS tumour genes by
combining real-time PCR with a hybridization technique. They show
that Eprobes provide decisive advantages over commonly used
hybridization probes because of their unique background signal
reduction, enhanced DNA-binding affinity and very low false
positives rate. The research is published in the journal PLOS ONE
“Eprobe enables real-time PCR methods, which are gaining
importance for medical diagnostics and many life science
applications, because they can provide quantitative results and
increased reliability as compared to standard PCR methods,” explains
Kengo Usui, the leader of the Genetic Diagnosis Technology Unit at
RIKEN Center for Life Science Technologies.
“This new technology will enable the development of advanced
assay formats for the simultaneous detection of multiple target
genes, as needed for example in the diagnosis of tumours” explains
Takeshi Hanami, first author of the paper.
“We are very excited about the potential of the new Eprobes”,
comments Matthias Harbers, Visiting Scientist to the Division of
Genomic Technologies at the RIKEN Center for Life Science
Technologies and supervisor of the Eprobe development project. “In
the reactions, Eprobes acted like sequence-specific fluorescent
dyes, which gives them great potential for use as hybridization
probes not only in PCR and melting curve analysis but also in other
important applications like for instance in Fluorescent in situ
Hybridization or FISH.”
1. Hanami et al. Eprobe mediated real-time PCR monitoring and
melting curve analysis. PLOS ONE, 2013.