Paar Christian, Hammerl Verena, Blessberger Hermann, Stekel Herbert, Steinwender Clemens, Berg Jörg
Clin Lab. 2016 Dec 1;62(12):2461-2467. doi: 10.7754/Clin.Lab.2016.160616.
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.
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.
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.
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.
扩增子的高分辨率熔解曲线分析（HRM）是一种用于单核苷酸多态性（SNP）基因分型的简单方法。尽管已有许多应用报道，但HRM似乎在临床实验室中很少使用。本研究对HRM-PCR在临床实验室中用于维生素K环氧化物还原酶复合体亚单位1基因SNP基因分型的适用性进行了调查。
在Cobas z480仪器上通过两种不同的HRM-PCR对约100份DNA样本进行分析，并与在LightCycler 2.0仪器上进行的荧光标记探针PCR（HybProbe-PCR）作为参考进行比较。
当扩增子大小较小（63 bp）且DNA输入量通过例如用无盐水稀释样本等方式受到限制时，可获得100%匹配结果的可靠基因分型。
通过不同方法提取的DNA可用于HRM-PCR基因分型。与HybProbe-PCR相比，Cobas z480仪器上的HRM-PCR具有更高的通量，然而，代价是更高程度的实验室标准化和更长的周转时间。
