Towards an integrated resource for the study of population and disease associated variability of the human mitochondrial genome

n/

Methodology for Y Chromosome Capture: A complete genome sequence of Y chromosome using flow cytometry, laser microdissection and magnetic streptavidin-beads

This study is a comparison of the efficiency of three technologies used for Y chromosome capture and the next-generation sequencing (NGS) technologies applied for determining its whole sequence. Our main findings disclose that streptavidin–biotin magnetic particle-based capture methodology offers better and a deeper sequence coverage for Y chromosome capture, compared to chromosome sorting and microdissection procedures. Moreover, this methodology is less time consuming and the most selective for capturing only Y chromosomal material, in contrast with other methodologies that result in considerable background material from other, non-targeted chromosomes. NGS results compared between two platforms, NextSeq 500 and SOLID 5500xl, produce the same coverage results. This is the first study to explore a methodological comparison of Y chromosome capture and genetic analysis. Our results indicate an improved strategy for Y chromosome research with applications in several scientific fields where this chromosome plays an important role, such as forensics, medical sciences, molecular anthropology and cancer sciences.Spanish Alfonso Martin Escudero Foundation for the financial support to one of the authors of the present work (MJ Alvarez –Cubero)

MToolBox: a highly authomated pipeline for heteroplasmy annotation and prioritization analysis of human mitochondrial variants in high-throughput sequencing

Abstract Motivation: The increasing availability of mitochondria-targeted and off-target sequencing data in whole-exome and whole-genome sequencing studies (WXS and WGS) has risen the demand of effective pipelines to accurately measure heteroplasmy and to easily recognize the most functionally important mitochondrial variants among a huge number of candidates. To this purpose, we developed MToolBox, a highly automated pipeline to reconstruct and analyze human mitochondrial DNA from high-throughput sequencing data. Results: MToolBox implements an effective computational strategy for mitochondrial genomes assembling and haplogroup assignment also including a prioritization analysis of detected variants. MToolBox provides a Variant Call Format file featuring, for the first time, allele-specific heteroplasmy and annotation files with prioritized variants. MToolBox was tested on simulated samples and applied on 1000 Genomes WXS datasets. Availability and implementation: MToolBox package is available at https://sourceforge.net/projects/mtoolbox/

High-Throughput Sequencing to Detect DNA-RNA Changes

: The advent of deep sequencing technologies has greatly improved the study of complex eukaryotic genomes and transcriptomes, allowing the investigation of posttranscriptional molecular mechanisms as alternative splicing and RNA editing at unprecedented throughput and resolution. The most prevalent type of RNA editing in higher eukaryotes is the deamination of adenosine to inosine (A-to-I) in double-stranded RNAs. Depending on the RNA type or the RNA region involved, A-to-I RNA editing contributes to the transcriptome and proteome diversity.Hereafter, we present an easy and reproducible computational protocol for the identification of candidate RNA editing sites in humans using deep transcriptome (RNA-Seq) and genome (DNA-Seq) sequencing

Mitochondrial Disease Sequence Data Resource (MSeqDR): A global grass-roots consortium to facilitate deposition, curation, annotation, and integrated analysis of genomic data for the mitochondrial disease clinical and research communities.

Abstract Success rates for genomic analyses of highly heterogeneous disorders can be greatly improved if a large cohort of patient data is assembled to enhance collective capabilities for accurate sequence variant annotation, analysis, and interpretation. Indeed, molecular diagnostics requires the establishment of robust data resources to enable data sharing that informs accurate understanding of genes, variants, and phenotypes. The "Mitochondrial Disease Sequence Data Resource (MSeqDR) Consortium" is a grass-roots effort facilitated by the United Mitochondrial Disease Foundation to identify and prioritize specific genomic data analysis needs of the global mitochondrial disease clinical and research community. A central Web portal (https://mseqdr.org) facilitates the coherent compilation, organization, annotation, and analysis of sequence data from both nuclear and mitochondrial genomes of individuals and families with suspected mitochondrial disease. This Web portal provides users with a flexible and expandable suite of resources to enable variant-, gene-, and exome-level sequence analysis in a secure, Web-based, and user-friendly fashion. Users can also elect to share data with other MSeqDR Consortium members, or even the general public, either by custom annotation tracks or through the use of a convenient distributed annotation system (DAS) mechanism. A range of data visualization and analysis tools are provided to facilitate user interrogation and understanding of genomic, and ultimately phenotypic, data of relevance to mitochondrial biology and disease. Currently available tools for nuclear and mitochondrial gene analyses include an MSeqDR GBrowse instance that hosts optimized mitochondrial disease and mitochondrial DNA (mtDNA) specific annotation tracks, as well as an MSeqDR locus-specific database (LSDB) that curates variant data on more than 1300 genes that have been implicated in mitochondrial disease and/or encode mitochondria-localized proteins. MSeqDR is integrated with a diverse array of mtDNA data analysis tools that are both freestanding and incorporated into an online exome-level dataset curation and analysis resource (GEM.app) that is being optimized to support needs of the MSeqDR community. In addition, MSeqDR supports mitochondrial disease phenotyping and ontology tools, and provides variant pathogenicity assessment features that enable community review, feedback, and integration with the public ClinVar variant annotation resource. A centralized Web-based informed consent process is being developed, with implementation of a Global Unique Identifier (GUID) system to integrate data deposited on a given individual from different sources. Community-based data deposition into MSeqDR has already begun. Future efforts will enhance capabilities to incorporate phenotypic data that enhance genomic data analyses. MSeqDR will fill the existing void in bioinformatics tools and centralized knowledge that are necessary to enable efficient nuclear and mtDNA genomic data interpretation by a range of shareholders across both clinical diagnostic and research settings. Ultimately, MSeqDR is focused on empowering the global mitochondrial disease community to better define and explore mitochondrial diseases