Protocol proposal for Friedreich ataxia molecular diagnosis using fluorescent and triplet repeat primed polymerase chain reaction

Xunclà, Mar; Rodríguez‐Revenga, Laia; Madrigal, Irene; Jiménez, Dolores; Milá, Montserrat; Badenas, Cèlia

doi:10.1016/j.trsl.2010.08.001

Cited by 7 publications

(8 citation statements)

References 9 publications

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“…Long-range PCR, which is highly effective at diagnosing homozygous GAA-TREs, has a high false-negative rate for heterozygous GAA-TRE. [14][15][16] Possible reasons include preferential PCR amplification of the normal allele (allele dropout) 14 and high levels of somatic instability of the GAA-TRE, 19 20 both of which are more likely with long GAA-TREs (>1000 triplets). Long-range PCR also has a false-positive rate, where a heteroduplex formed between normal alleles of variable length Neurogenetics masquerades as a GAA-TRE.…”

Section: Discussionmentioning

confidence: 99%

“…18 In a cohort of 310 individuals (with 38 homozygotes and 55 heterozygous carriers), long-range PCR mischaracterised 25 individuals; 14 heterozygous carriers were identified as not having a GAA-TRE, and 11 individuals without a GAA-TRE were identified as being heterozygous carriers. 14 TP-PCR is efficient at detecting the presence of an expanded allele, [14][15][16] although it typically finds that the GAA-TRE is above a certain threshold length without providing an accurate estimate of the length of the GAA-TRE. Southern blot, while accurate at detecting GAA-TREs in the heterozygous state, is tedious and requires large quantities of DNA.…”

Section: Discussionmentioning

confidence: 99%

“…14-16 18 Some laboratories use protocols that incorporate more than one technique in order to avoid misdiagnosing heterozygous carriers. [14][15][16] Assuming that heterozygous carriers of GAA-TREs should have ~50% methylation in the FRDA-DMR, 8 we explored if this epigenetic readout could effectively identify GAA-TREs in the heterozygous state. In a series of known heterozygous carriers of the GAA-TRE, we found that methylation of the FRDA-DMR is a reliable way to identify heterozygosity.…”

Section: Introductionmentioning

confidence: 99%

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FXNgene methylation determines carrier status in Friedreich ataxia

et al. 2023

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BackgroundFriedreich ataxia (FRDA) is typically caused by homozygosity for an expanded GAA triplet-repeat (GAA-TRE) in intron 1 of theFXNgene. Some patients are compound heterozygous for the GAA-TRE and anotherFXNpathogenic variant. Detection of the GAA-TRE in the heterozygous state, occasionally technically challenging, is essential for diagnosing compound heterozygotes and asymptomatic carriers.ObjectiveWe explored if the FRDA differentially methylated region (FRDA-DMR) in intron 1, which is hypermethylated inciswith the GAA-TRE, effectively detects heterozygous GAA-TRE.MethodsFXNDNA methylation was assayed by targeted bisulfite deep sequencing using the Illumina platform.ResultsFRDA-DMR methylation effectively identified a cohort of known heterozygous carriers of the GAA-TRE. In an individual with clinical features of FRDA, commercial testing showed a paternally inherited pathogenicFXNinitiation codon variant but no GAA-TRE. Methylation in the FRDA-DMR effectively identified the proband, his mother and various maternal relatives as heterozygous carriers of the GAA-TRE, thus confirming the diagnosis of FRDA.ConclusionFXNDNA methylation reliably detects the GAA-TRE in the heterozygous state and offers a robust alternative strategy to diagnose FRDA due to compound heterozygosity and to identify asymptomatic heterozygous carriers of the GAA-TRE.

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Section: Discussionmentioning

confidence: 99%

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

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FXNgene methylation determines carrier status in Friedreich ataxia

et al. 2023

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“…2C). In this situation, further molecular tests are needed, such as analysis of FXN exons (1-5a and 5b) for point mutations [19,41] [42] and for exonic deletions [20].…”

Section: Pathogenesis and Molecular Basismentioning

confidence: 99%

“…This method showed highly reliable and sensitive results, and it is a robust technique which overcomes the shortcomings of a traditional PCR by detecting the very large expanded alleles of even >5 kb accurately [42][43][44].…”

Section: Pathogenesis and Molecular Basismentioning

confidence: 99%

Milestones in Friedreich ataxia: more than a century and still learning

et al. 2015

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Friedreich ataxia (FRDA) is the most common autosomal recessive ataxia worldwide. This review highlights the main clinical features, pathophysiological mechanisms, and therapeutic approaches for FRDA patients. The disease is characterized by a combination of neurological involvement (ataxia and neuropathy), cardiomyopathy, skeletal abnormalities, and glucose metabolism disturbances. FRDA is caused by expanded guanine-adenine-adenine (GAA) triplet repeats in the first intron of the frataxin gene (FXN), resulting in reduction of messenger RNA and protein levels of frataxin in different tissues. The molecular and metabolic disturbances, including iron accumulation, lead to pathological changes characterized by spinal cord and dorsal root ganglia atrophy, dentate nucleus atrophy without global cerebellar volume reduction, and hypertrophic cardiomyopathy. DNA analysis is the hallmark for the diagnosis of FRDA. There is no specific treatment to stop the disease progression in FRDA patients. However, a number of drugs are under investigation. Therapeutic approaches intend to improve mitochondrial functioning and to increase FXN expression.

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Hereditary Myelopathy: A Clinical Approach

Fink

2022

Myelopathy

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Protocol proposal for Friedreich ataxia molecular diagnosis using fluorescent and triplet repeat primed polymerase chain reaction

Cited by 7 publications

References 9 publications

FXNgene methylation determines carrier status in Friedreich ataxia

FXNgene methylation determines carrier status in Friedreich ataxia

Milestones in Friedreich ataxia: more than a century and still learning

Hereditary Myelopathy: A Clinical Approach

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