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Genetic Variant Detection in the CALR gene using High Resolution Melting Analysis
Published on: August 26, 2020
Calcium-sensing receptor mutations and denaturing high performance liquid chromatography
David E C Cole1, Francisco H J Yun, Betty Y L Wong
1Departments of Laboratory Medicine and Pathobiology, Medicine, and Genetics, University of Toronto, Ontario, Canada.
Journal of Molecular Endocrinology
|January 31, 2009
Summary
A new high-throughput method using denaturing high performance liquid chromatography (DHPLC) rapidly screens for calcium-sensing receptor (CASR) mutations. This technique accurately identifies mutations linked to familial hypocalciuric hypercalcemia (FHH) and autosomal dominant hypocalcemia (ADH).
Area of Science:
- Biochemistry
- Genetics
- Molecular Biology
Background:
- The calcium-sensing receptor (CASR) regulates calcium homeostasis and is crucial in parathyroid and kidney function.
- CASR mutations cause disorders like familial hypocalciuric hypercalcemia (FHH) and autosomal dominant hypocalcemia (ADH).
- Accurate identification of CASR mutations is vital for managing mineral metabolism disorders.
Purpose of the Study:
- To develop and validate a high-throughput screening method for CASR mutations.
- To efficiently detect novel and recurrent mutations associated with FHH and ADH.
Main Methods:
- Utilized denaturing high performance liquid chromatography (DHPLC) for high-throughput screening of 13 CASR amplicons.
- Employed sequencing for amplicons with altered melting curves and direct sequencing for the COOH-terminal tail.
- Conducted a blinded analysis using 32 positive and 22 negative controls.
Main Results:
- Achieved 100% concordance in the blinded analysis.
- Identified eight novel and five recurrent FHH-associated mutations.
- Detected six novel and two recurrent ADH-associated mutations.
Conclusions:
- DHPLC offers a rapid and effective method for screening CASR mutations.
- This approach aids in the clinical diagnosis and management of calcium-related disorders.
- The study identified numerous new and recurring mutations in CASR.
