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Updated: Jun 25, 2025

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Establishing a Mouse Genotyping Core Facility Based on Automation, High-resolution Melting Analysis, and

Hagit Dafni1, Kohava Levi1, Miriam Chen Eldad1

  • 1Department of Veterinary Resources, The Weizmann Institute of Science, Rehovot, Israel.

Comparative Medicine
|May 25, 2024
PubMed
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Establishing an in-house genotyping facility using automated PCR assembly and high-resolution melting (HRM) analysis improves cost-effectiveness and reliability. This refined process ensures accurate mouse colony management and reproducible research outcomes.

Area of Science:

  • Molecular Biology
  • Genetics
  • Laboratory Management

Background:

  • Genotyping is crucial for efficient mouse colony management and research reproducibility.
  • Outsourced genotyping services can be costly, and in-house methods using temporary staff lack reliability.
  • Establishing a dedicated, reliable, and cost-effective in-house genotyping facility is essential for research institutions.

Purpose of the Study:

  • To describe the establishment of an in-house genotyping core facility.
  • To implement automated Polymerase Chain Reaction (PCR) assembly and high-resolution melting (HRM) analysis for enhanced genotyping.
  • To develop a streamlined workflow for efficient sample tracking, data analysis, and report generation.

Main Methods:

  • Utilized automated PCR assembly and high-resolution melting (HRM) analysis for genotyping.

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  • Developed custom MATLAB applications for workflow management, sample tracking, and data analysis.
  • Transitioned analysis of over 250 target genes from gel electrophoresis to HRM analysis.

    • Main Results:

    • Successfully transitioned over 250 target genes to HRM analysis, with only 2% requiring protocol adjustments.
    • Achieved high throughput, processing over 1,000 PCR reactions weekly.
    • Demonstrated that blastocyst genotyping can serve as quality control for cryopreservation, supporting the 3Rs principles.

    Conclusions:

    • The developed in-house genotyping protocol is cost-effective, reliable, and efficient.
    • The automated workflow and HRM analysis enhance accuracy and reproducibility in research.
    • The protocol supports the 3Rs principles (Replacement, Reduction, and Refinement) in animal research.