A rapid detection method for the ryanodine receptor 1 (C7360G) mutation in quarter horses

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Abstract

Background: Anesthetic-induced malignant hyperthermia has been documented in Quarter Horses and is caused by a single-point mutation in the ryanodine receptor 1 gene at nucleotide C7360G generating a R2454G amino acid substitution. An accurate, faster molecular test that is less prone to contamination would facilitate screening for the mutation in horses intended for breeding, in those undergoing surgical procedures, and in those with clinical signs compatible with malignant hyperthermia. Objective: To report a rapid and accurate method for the detection of the ryanodine receptor 1 C7360G mutation. Animals: Eleven diseased, 10 healthy, and 225 randomly selected Quarter Horses. Methods: This study included horses with the ryanodine receptor 1 C7360G mutation as detected by gene sequencing. Available genomic and complementary DNA extracted from whole blood, hair or skeletal muscle was used for genetic analysis. Real-time polymerase chain reaction (RT-PCR) melting curve analysis was performed by equine specific primers and 2 hybridization probes (sensor and anchor probes) that contain the site of the mutation. Results from this method were blinded and compared with nucleic acid sequencing for validation. Results: A rapid genotyping assay with fluorescence resonance energy transfer probes and melting curve analysis was accurate (100% agreement, K 5 1) for identification of affected horses. The prevalence of the mutation in a random population of Quarter Horses was 1.3%. Conclusions and Clinical Importance: Malignant hyperthermia in Quarter Horses can be rapidly and accurately detected by RT-PCR melting curve genotyping with hybridization probes.

Original languageEnglish (US)
Pages (from-to)619-622
Number of pages4
JournalJournal of Veterinary Internal Medicine
Volume23
Issue number3
DOIs
StatePublished - May 2009

Fingerprint

ryanodine receptors
Quarter Horse
Ryanodine Receptor Calcium Release Channel
Horses
malignant hyperthermia
mutation
Mutation
melting
Malignant Hyperthermia
horses
genotyping
Freezing
quantitative polymerase chain reaction
methodology
complementary DNA
livestock breeding
Real-Time Polymerase Chain Reaction
amino acid substitution
point mutation
energy transfer

Keywords

  • Equine
  • Malignant hyperthermia
  • Melting curve genotyping

ASJC Scopus subject areas

  • veterinary(all)

Cite this

@article{acddf85791974b17bea048ec6d38f136,
title = "A rapid detection method for the ryanodine receptor 1 (C7360G) mutation in quarter horses",
abstract = "Background: Anesthetic-induced malignant hyperthermia has been documented in Quarter Horses and is caused by a single-point mutation in the ryanodine receptor 1 gene at nucleotide C7360G generating a R2454G amino acid substitution. An accurate, faster molecular test that is less prone to contamination would facilitate screening for the mutation in horses intended for breeding, in those undergoing surgical procedures, and in those with clinical signs compatible with malignant hyperthermia. Objective: To report a rapid and accurate method for the detection of the ryanodine receptor 1 C7360G mutation. Animals: Eleven diseased, 10 healthy, and 225 randomly selected Quarter Horses. Methods: This study included horses with the ryanodine receptor 1 C7360G mutation as detected by gene sequencing. Available genomic and complementary DNA extracted from whole blood, hair or skeletal muscle was used for genetic analysis. Real-time polymerase chain reaction (RT-PCR) melting curve analysis was performed by equine specific primers and 2 hybridization probes (sensor and anchor probes) that contain the site of the mutation. Results from this method were blinded and compared with nucleic acid sequencing for validation. Results: A rapid genotyping assay with fluorescence resonance energy transfer probes and melting curve analysis was accurate (100{\%} agreement, K 5 1) for identification of affected horses. The prevalence of the mutation in a random population of Quarter Horses was 1.3{\%}. Conclusions and Clinical Importance: Malignant hyperthermia in Quarter Horses can be rapidly and accurately detected by RT-PCR melting curve genotyping with hybridization probes.",
keywords = "Equine, Malignant hyperthermia, Melting curve genotyping",
author = "Jorge Nieto and Aleman, {Monica R}",
year = "2009",
month = "5",
doi = "10.1111/j.1939-1676.2009.0281.x",
language = "English (US)",
volume = "23",
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journal = "Journal of Veterinary Internal Medicine",
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T1 - A rapid detection method for the ryanodine receptor 1 (C7360G) mutation in quarter horses

AU - Nieto, Jorge

AU - Aleman, Monica R

PY - 2009/5

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N2 - Background: Anesthetic-induced malignant hyperthermia has been documented in Quarter Horses and is caused by a single-point mutation in the ryanodine receptor 1 gene at nucleotide C7360G generating a R2454G amino acid substitution. An accurate, faster molecular test that is less prone to contamination would facilitate screening for the mutation in horses intended for breeding, in those undergoing surgical procedures, and in those with clinical signs compatible with malignant hyperthermia. Objective: To report a rapid and accurate method for the detection of the ryanodine receptor 1 C7360G mutation. Animals: Eleven diseased, 10 healthy, and 225 randomly selected Quarter Horses. Methods: This study included horses with the ryanodine receptor 1 C7360G mutation as detected by gene sequencing. Available genomic and complementary DNA extracted from whole blood, hair or skeletal muscle was used for genetic analysis. Real-time polymerase chain reaction (RT-PCR) melting curve analysis was performed by equine specific primers and 2 hybridization probes (sensor and anchor probes) that contain the site of the mutation. Results from this method were blinded and compared with nucleic acid sequencing for validation. Results: A rapid genotyping assay with fluorescence resonance energy transfer probes and melting curve analysis was accurate (100% agreement, K 5 1) for identification of affected horses. The prevalence of the mutation in a random population of Quarter Horses was 1.3%. Conclusions and Clinical Importance: Malignant hyperthermia in Quarter Horses can be rapidly and accurately detected by RT-PCR melting curve genotyping with hybridization probes.

AB - Background: Anesthetic-induced malignant hyperthermia has been documented in Quarter Horses and is caused by a single-point mutation in the ryanodine receptor 1 gene at nucleotide C7360G generating a R2454G amino acid substitution. An accurate, faster molecular test that is less prone to contamination would facilitate screening for the mutation in horses intended for breeding, in those undergoing surgical procedures, and in those with clinical signs compatible with malignant hyperthermia. Objective: To report a rapid and accurate method for the detection of the ryanodine receptor 1 C7360G mutation. Animals: Eleven diseased, 10 healthy, and 225 randomly selected Quarter Horses. Methods: This study included horses with the ryanodine receptor 1 C7360G mutation as detected by gene sequencing. Available genomic and complementary DNA extracted from whole blood, hair or skeletal muscle was used for genetic analysis. Real-time polymerase chain reaction (RT-PCR) melting curve analysis was performed by equine specific primers and 2 hybridization probes (sensor and anchor probes) that contain the site of the mutation. Results from this method were blinded and compared with nucleic acid sequencing for validation. Results: A rapid genotyping assay with fluorescence resonance energy transfer probes and melting curve analysis was accurate (100% agreement, K 5 1) for identification of affected horses. The prevalence of the mutation in a random population of Quarter Horses was 1.3%. Conclusions and Clinical Importance: Malignant hyperthermia in Quarter Horses can be rapidly and accurately detected by RT-PCR melting curve genotyping with hybridization probes.

KW - Equine

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