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Updated: Mar 7, 2026
Rapid and Efficient Zebrafish Genotyping Using PCR with High-resolution Melt Analysis
Published on: February 5, 2014
Conditions of High Resolution Melting Analysis on the Cobas z480 Instrument for the Genotyping of VKORC1 in the
Background:
High resolution melting (HRM) of amplicons is a simple method for genotyping of single nucleotide polymorphisms (SNPs). Albeit many applications reported, HRM seems to be rarely used in clinical laboratories. The suitability of HRM-PCR for the clinical laboratory was investigated for genotyping of SNPs of the vitamin K epoxide reductase complex unit 1 gene.
Methods:
About 100 DNA samples were analyzed by two different HRM-PCRs on the Cobas z480 instrument and compared with a PCR with fluorescently labeled probes (HybProbe-PCR) on the LightCycler 2.0 instrument as reference.
Results:
Reliable genotyping with 100% matching results was obtained, when the amplicon size was small (63 bp) and DNA input was limited by e.g., sample dilution with salt-free water.
Conclusions:
DNA extracted by differing methods may be used for genotyping by HRM-PCR. Compared with HybProbe-PCR, HRM-PCR on the Cobas z480 instrument allows for higher through-put, however, at the cost of a higher degree of laboratory standardization and a slower turnaround.
