Molecular fingerprints to identify Candida species

Abstract

A wide range of molecular techniques have been developed for genotyping Candida species. Among them, multilocus sequence typing (MLST) and microsatellite length polymorphisms (MLP) analysis have recently emerged. MLST relies on DNA sequences of internal regions of various independent housekeeping genes, while MLP identifies microsatellite instability. Both methods generate unambiguous and highly reproducible data. Here, we review the results achieved by using these two techniques and also provide a brief overview of a new method based on high-resolution DNA melting (HRM). This method identifies sequence differences by subtle deviations in sample melting profiles in the presence of saturating fluorescent DNA binding dyes.

Figure 1

Schematic representation of HRM analysis for SNPs genotyping. Arrows indicate the positions of the primers for allele amplification of a region harboring a SNP. The DNA fluorophore has a bright fluorescence when intercalated to double-stranded DNA (black circle) and low fluorescence in the unbound state (gray circles). Mispaired nucleotides are shown as diagonally broken lines. PCR products from homozygous wild type (solid lines), heterozygous mutant (dotted lines), and homozygous mutant (dashed lines) were analyzed by normalized melting curves (a), derivate melting curves (b), and difference plots (c).