Finally, we have demonstrated a 100% concordance between the in vivo and in vitro porcine stress syndrome (PSS) susceptibility results and the ones obtained by means of molecular genotyping such as real-time PCR using fluorescein-labeled hybridization probe
, sequencing and restriction analysis.
Acknowledgments: We thank all the patients and their families for their kind cooperation; Jan Schouten and his team at MRC-Holland for providing hybridization probe
The sequences of additional primers and dual hybridization probes
for BRAF p.V600E have previously been published (27).
Using the nearest neighbor model for the estimation of matched and mismatched hybridization probe
melting points and selection of optimal probes on the light-cycler.
The hybridization probes
contain a 5' MGB and fluorophore with a 3' quencher.
The methods used for this genotyping were LightCycler (Roche Applied Science, Mannheim, Germany) real-time PCR and fluorescence resonance energy transfer hybridization probes
In the hybridization probe
format, the rapid loss of fluorescence is observed when the temperature reaches the Tm of the probes.
Andreas Nitsche for designing TBP primers and hybridization probe
Because melting curve analysis with hybridization probe
technology is routinely used in our laboratory (3), we conducted a study to implement IVS8 genotyping and confirm its reliability.
Application of a thermodynamic nearest-neighbor model to estimate nucleic acid stability and optimize probe design: prediction of melting points of multiple mutations of apolipoprotein B-3500 and factor V with a hybridization probe
genotyping assay on the LightCycler.
For each RET exon, sample DNA (~50 ng total) was amplified with the LightCycler[R] FastStart DNA Master Hybridization Probe
Kit (Roche Diagnostics Corp.) in a final PCR reaction volume of 10 [micro]L.
The product portfolio includes Fluorescence in Situ Hybridization and Chromogenic in Situ Hybridization probes
that are designed and optimized for specific diseases, genes, and/or regions across the entire human or mouse genomes.