*5.3.9 High resolution melt curve (HRM)*

High resolution melt curve typically involve qPCR amplification followed by a melting curve collected using a fluorescent dye. It follows extraction of nucleic acids and qPCR with fluorescent labeled dye. Following amplification and quantification of the 16S rRNA gene, a high-resolution melting curve analysis is obtained. The procedure melts the amplified 16S rRNA gene products between the temperatures ranging from 72°C and ending at 95°C, with fluorescence measurements taken at every 0.1°C increment. Melting curves are normalized to relative fluorescence units (RFU) in a specified "melt region" (e.g., 83.5–89.5°C), thereby negating the effect of absolute RFU values. The melt region is autocalled by the melt analysis software (Precision Melt Analysis) [61]. The effects of herbicide on soil bacterial diversity were efficiently studied using HRM analysis [62].

*Soil Metagenomics: Concepts and Applications DOI: http://dx.doi.org/10.5772/intechopen.88958*
