Work

Protocol for detection and typing of Serratia marcescens using a qPCR-based technique without cultivation or sequencing

Year: 2025

Type: article

Abstract: High-resolution melting (HRM) qPCR is a molecular biology technique for quantifying DNA targets and discriminating them on the basis of their sequences. Hypervariable-locus melting typing (HLMT) exploits HRM for bacterial typing by targeting variable genetic loci. Here, we present an HLMT protocol for detecting Serratia marcescens clones directly from biological samples without cultivation. We describe steps for DNA extraction, endpoint PCR, and HRM qPCR. We then detail procedures for data analysis and interpretation. For complete details on the use and execution of this protocol, please refer to Alvaro et al. 1 • Guidance on detecting and typing Serratia marcescens directly from samples without cultivation • Steps for HRM qPCR on unique hypervariable loci , compatible with standard qPCR instruments • Procedures for rapid, cost-effective typing, for outbreak control and large screenings Publisher’s note: Undertaking any experimental protocol requires adherence to local institutional guidelines for laboratory safety and ethics. High-resolution melting (HRM) qPCR is a molecular biology technique for quantifying DNA targets and discriminating them on the basis of their sequences. Hypervariable-locus melting typing (HLMT) exploits HRM for bacterial typing by targeting variable genetic loci. Here, we present an HLMT protocol for detecting Serratia marcescens clones directly from biological samples without cultivation. We describe steps for DNA extraction, endpoint PCR, and HRM qPCR. We then detail procedures for data analysis and interpretation.

Source: STAR Protocols

DOI: 10.1016/j.xpro.2025.104216