Mitochondrial DNA analysis by multiplex denaturing high-performance liquid chromatography and selective sequencing in pediatric patients with cardiomyopathy

Schrijver, Iris; Pique, Lynn; Traynis, Ilana; Scharfe, Curt; Sehnert, Amy J.

doi:10.1097/gim.0b013e318190356b

Cited by 6 publications

(8 citation statements)

References 34 publications

Supporting

Mentioning

Contrasting

Order By: Relevance

“…The approach used two additional factors, haplogroup and allele frequencies, along with standard criteria to evaluate the long list of mtDNA variants detected in molecular studies on cardiomyopathy. 6, 7, 8, 15, 16, 17 To access large numbers of reference mtDNAs, we used online resources such as GenBank. Using this system, we were able to eliminate 98% of the 270 different variants and excluded 79% of the cases (23 of 29).…”

Section: Discussionmentioning

confidence: 99%

“…6, 7, 8, 15, 16, 17 The mtDNAs of over 500 patients with HCM or DCM patients have already been sequenced and the mtDNA variants that differ from a reference sequence have been compared with small samples of ‘control' cases. Those variants observed in the patient mtDNAs but not in the study controls have been considered to be potential disease causing mutations.…”

Section: Introductionmentioning

confidence: 99%

“…Over 1000 variants have been reported in various cardiomyopathy cases, which encompass 200 different sequence variants. 6, 7, 8, 15, 16, 17 …”

Section: Introductionmentioning

confidence: 99%

See 2 more Smart Citations

Mitochondrial cardiomyopathies: how to identify candidate pathogenic mutations by mitochondrial DNA sequencing, MITOMASTER and phylogeny

et al. 2010

View full text Add to dashboard Cite

Pathogenic mitochondrial DNA (mtDNA) mutations leading to mitochondrial dysfunction can cause cardiomyopathy and heart failure. Owing to a high mutation rate, mtDNA defects may occur at any nucleotide in its 16 569 bp sequence. Complete mtDNA sequencing may detect pathogenic mutations, which can be difficult to interpret because of normal ethnic/geographic-associated haplogroup variation. Our goal is to show how to identify candidate mtDNA mutations by sorting out polymorphisms using readily available online tools. The purpose of this approach is to help investigators in prioritizing mtDNA variants for functional analysis to establish pathogenicity. We analyzed complete mtDNA sequences from 29 Italian patients with mitochondrial cardiomyopathy or suspected disease. Using MITOMASTER and PhyloTree, we characterized 593 substitution variants by haplogroup and allele frequencies to identify all novel, non-haplogroup-associated variants. MITOMASTER permitted determination of each variant's location, amino acid change and evolutionary conservation. We found that 98% of variants were common or rare, haplogroup-associated variants, and thus unlikely to be primary cause in 80% of cases. Six variants were novel, non-haplogroup variants and thus possible contributors to disease etiology. Two with the greatest pathogenic potential were heteroplasmic, nonsynonymous variants: m.15132T>C in MT-CYB for a patient with hypertrophic dilated cardiomyopathy and m.6570G>T in MT-CO1 for a patient with myopathy. In summary, we have used our automated information system, MITOMASTER, to make a preliminary distinction between normal mtDNA variation and pathogenic mutations in patient samples; this fast and easy approach allowed us to select the variants for traditional analysis to establish pathogenicity.

show abstract

Section: Discussionmentioning

confidence: 99%

Section: Introductionmentioning

confidence: 99%

See 1 more Smart Citation

Mitochondrial cardiomyopathies: how to identify candidate pathogenic mutations by mitochondrial DNA sequencing, MITOMASTER and phylogeny

et al. 2010

View full text Add to dashboard Cite

show abstract

“…Traditionally, the most comprehensive method used to detect variants is Sanger capillary sequencing of whole mtDNA [8], [9]. However, limitations prevent the routine use of whole mtDNA studies by Sanger sequencing including the amount of labor and time to manually inspect data for heteroplasmic mutations.…”

Section: Introductionmentioning

confidence: 99%

Mitochondrial DNA Variant Discovery and Evaluation in Human Cardiomyopathies through Next-Generation Sequencing

et al. 2010

View full text Add to dashboard Cite

Mutations in mitochondrial DNA (mtDNA) may cause maternally-inherited cardiomyopathy and heart failure. In homoplasmy all mtDNA copies contain the mutation. In heteroplasmy there is a mixture of normal and mutant copies of mtDNA. The clinical phenotype of an affected individual depends on the type of genetic defect and the ratios of mutant and normal mtDNA in affected tissues. We aimed at determining the sensitivity of next-generation sequencing compared to Sanger sequencing for mutation detection in patients with mitochondrial cardiomyopathy. We studied 18 patients with mitochondrial cardiomyopathy and two with suspected mitochondrial disease. We “shotgun” sequenced PCR-amplified mtDNA and multiplexed using a single run on Roche's 454 Genome Sequencer. By mapping to the reference sequence, we obtained 1,300× average coverage per case and identified high-confidence variants. By comparing these to >400 mtDNA substitution variants detected by Sanger, we found 98% concordance in variant detection. Simulation studies showed that >95% of the homoplasmic variants were detected at a minimum sequence coverage of 20× while heteroplasmic variants required >200× coverage. Several Sanger “misses” were detected by 454 sequencing. These included the novel heteroplasmic 7501T>C in tRNA serine 1 in a patient with sudden cardiac death. These results support a potential role of next-generation sequencing in the discovery of novel mtDNA variants with heteroplasmy below the level reliably detected with Sanger sequencing. We hope that this will assist in the identification of mtDNA mutations and key genetic determinants for cardiomyopathy and mitochondrial disease.

show abstract

“…On the contrary, we detected in patient B the m.14766G4A in the cytochrome B gene and the m.13708G4A in the MT-ND5 gene that was detected in patient with idiopathic cardiomyopathy (Schrijver et al, 2009). These two variations have been associated with Leber's hereditary optic neuropathy (LHON) especially the m.13708G4A which was considered as secondary mutation (Obermaier-Kusser et al, 1994;Oostra et al, 1994).…”

Section: Discussionmentioning

confidence: 81%

Whole mitochondrial genome analysis in two families with dilated mitochondrial cardiomyopathy: detection of mutations in MT-ND2 and MT-TL1 genes

Alila

Rebai

Tabebi

et al. 2015

Mitochondrial DNA Part A

View full text Add to dashboard Cite

Pathogenic mitochondrial DNA (mtDNA) mutations leading to mitochondrial dysfunction can cause cardiomyopathy and heart failure. These mutations were described in the mt-tRNA genes and in the mitochondrial protein-coding genes. The aim of this study was to identify the genetic defect in two patients belonging to two families with cardiac dysfunction associated to a wide spectrum of clinical phenotypes. The sequencing analysis of the whole mitochondrial DNA in the two patients and their parents revealed the presence of known polymorphisms associated to cardiomyopathy and two pathogenic mutations in DNA extracted from blood leucocytes: the heteroplasmic m.3243A > G mutation in the MT-TL1 gene in patient A; and the homoplasmic m.5182C > T mutation in the ND2 gene in patient B. Secondary structure analysis of the ND2 protein further supported the deleterious role of the m.5182C > T mutation, as it was found to be involved an extended imbalance in its hydrophobicity and affect its function. In addition, the mitochondrial variants identified in patients A and B classify both of them in the same haplogroup H2a2a1.

show abstract

Mitochondrial DNA analysis by multiplex denaturing high-performance liquid chromatography and selective sequencing in pediatric patients with cardiomyopathy

Cited by 6 publications

References 34 publications

Mitochondrial cardiomyopathies: how to identify candidate pathogenic mutations by mitochondrial DNA sequencing, MITOMASTER and phylogeny

Mitochondrial cardiomyopathies: how to identify candidate pathogenic mutations by mitochondrial DNA sequencing, MITOMASTER and phylogeny

Mitochondrial DNA Variant Discovery and Evaluation in Human Cardiomyopathies through Next-Generation Sequencing

Whole mitochondrial genome analysis in two families with dilated mitochondrial cardiomyopathy: detection of mutations in MT-ND2 and MT-TL1 genes

Contact Info

Product

Resources

About