Dataset Information

Mitochondrial and nuclear disease panel (Mito-aND-Panel): Combined sequencing of mitochondrial and nuclear DNA by a cost-effective and sensitive NGS-based method.

ABSTRACT: BACKGROUND:The diagnosis of mitochondrial disorders is challenging because of the clinical variability and genetic heterogeneity of these conditions. Next-Generation Sequencing (NGS) technology offers a robust high-throughput platform for nuclear and mitochondrial DNA (mtDNA) analyses. METHOD:We developed a custom Agilent SureSelect Mitochondrial and Nuclear Disease Panel (Mito-aND-Panel) capture kit that allows parallel enrichment for subsequent NGS-based sequence analysis of nuclear mitochondrial disease-related genes and the complete mtDNA genome. Sequencing of enriched mtDNA simultaneously with nuclear genes was compared with the separated sequencing of the mitochondrial genome and whole exome sequencing (WES). RESULTS:The Mito-aND-Panel permits accurate detection of low-level mtDNA heteroplasmy due to a very high sequencing depth compared to standard diagnostic procedures using Sanger sequencing/SNaPshot and WES which is crucial to identify maternally inherited mitochondrial disorders. CONCLUSION:We established a NGS-based method with combined sequencing of the complete mtDNA and nuclear genes which enables a more sensitive heteroplasmy detection of mtDNA mutations compared to traditional methods. Because the method promotes the analysis of mtDNA variants in large cohorts, it is cost-effective and simple to setup, we anticipate this is a highly relevant method for sequence-based genetic diagnosis in clinical diagnostic applications.

SUBMITTER: Abicht A

PROVIDER: S-EPMC6305657 | biostudies-literature | 2018 Nov

REPOSITORIES: biostudies-literature

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Publications

Mitochondrial and nuclear disease panel (Mito-aND-Panel): Combined sequencing of mitochondrial and nuclear DNA by a cost-effective and sensitive NGS-based method.

Abicht Angela A Scharf Florentine F Kleinle Stephanie S Schön Ulrike U Holinski-Feder Elke E Horvath Rita R Benet-Pagès Anna A Diebold Isabel I

Molecular genetics & genomic medicine 20181108 6

<h4>Background</h4>The diagnosis of mitochondrial disorders is challenging because of the clinical variability and genetic heterogeneity of these conditions. Next-Generation Sequencing (NGS) technology offers a robust high-throughput platform for nuclear and mitochondrial DNA (mtDNA) analyses.<h4>Method</h4>We developed a custom Agilent SureSelect Mitochondrial and Nuclear Disease Panel (Mito-aND-Panel) capture kit that allows parallel enrichment for subsequent NGS-based sequence analysis of nuc ...[more]

PMID: 30406974

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Project description:An increasing number of studies show that genetic markers can aid in refining prognostic information and predicting the benefit from systemic therapy. Our goal was to develop a high throughput, cost-effective and simple methodology for the detection of clinically relevant hot spot mutations in colon cancer.The Maldi-Tof mass spectrometry platform and OncoCarta panel from Sequenom were used to profile 239 colon cancers and 39 metastatic lymph nodes from NSABP clinical trial C-07 utilizing routinely processed FFPET (formalin-fixed paraffin-embedded tissue).Among the 238 common hot-spot cancer mutations in 19 genes interrogated by the OncoCarta panel, mutations were detected in 7 different genes at 26 different nucleotide positions in our colon cancer samples. Twenty-four assays that detected mutations in more than 1% of the samples were reconfigured into a new multiplexed panel, termed here as ColoCarta. Mutation profiling was repeated on 32 mutant samples using ColoCarta and the results were identical to results with OncoCarta, demonstrating that this methodology was reproducible. Further evidence demonstrating the validity of the data was the fact that the mutation frequencies of the most common colon cancer mutations were similar to the COSMIC (Catalog of Somatic Mutations in Cancer) database. The frequencies were 43.5% for KRAS, 20.1% for PIK3CA, and 12.1% for BRAF. In addition, infrequent mutations in NRAS, AKT1, ABL1, and MET were detected. Mutation profiling of metastatic lymph nodes and their corresponding primary tumors showed that they were 89.7% concordant. All mutations found in the lymph nodes were also found in the corresponding primary tumors, but in 4 cases a mutation was present in the primary tumor only.This study describes a high throughput technology that can be used to interrogate DNAs isolated from routinely processed FFPET and identifies the specific mutations that are common to colon cancer. The development of this technology and the ColoCarta panel may provide a mechanism for rapid screening of mutations in clinically relevant genes like KRAS, PIK3CA, and BRAF.ClinicalTrials.gov: NSABP C-07: NCT00004931.

Dataset Information

Mitochondrial and nuclear disease panel (Mito-aND-Panel): Combined sequencing of mitochondrial and nuclear DNA by a cost-effective and sensitive NGS-based method.

Publications

Mitochondrial and nuclear disease panel (Mito-aND-Panel): Combined sequencing of mitochondrial and nuclear DNA by a cost-effective and sensitive NGS-based method.

Similar Datasets

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