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Pipeline publication

[…] ranges were obtained using the provided standard immunoglobulin., DNA was extracted from EDTA blood using Maxwell 16 Blood DNA Purification Kit (Promega) or PBMC using DNeasy Blood & Tissue Kit (Qiagen). Total of 3 ug of DNA were sheared using E220 focused sonicator (Covaris) and exome libraries were generated using the SureSelect Human All Exon Kits (Agilent) according to manufactures’ protocol. The quality of generated libraries was inspected using Agilent High Sensitivity DNA Kit (Agilent) and quantified using qPCR kit (Agilent). Samples were sequenced on Illumina HiSeq2000 (Illumina) generating 100 bp paired end reads. Sequences were aligned to a human reference genome GRCh37 using bwa v 0.6.1 with default parameters. Variant calling (Single nucleotide variants and indels) was performed using GATK v.2 and variants were annotated using Annovar. An in-house custom analysis pipeline was used to filter and prioritize variants based on the likely genetic models and clinical pedigree for patients., DNA samples were extracted from blood or saliva using Maxwell 16 Blood DNA Purification Kit (Promega) and Oragene DNA (OG500) (Oragene), respectively. The candidate mutations in affected and unaffected individuals of both families were validated using BigDye Terminator Sequencing kit (Life technologies) and sequenced on ABI3730xl genetic analyser (Applied Biosystems). PCR primer sequences are available on request., All flow cytometry was carried out in a final staining volume of 100–200 μL, with data acquisition on an LSR II, LSRFort […]

Pipeline specifications

Software tools BWA, GATK, ANNOVAR
Organisms Mus musculus
Diseases Autoimmune Diseases, Disease, Blood Protein Disorders, Dysgammaglobulinemia, Immunologic Deficiency Syndromes, Inflammation, Genetic Diseases, Inborn
Chemicals Nucleotides