Computational protocol: Agammaglobulinaemia despite terminal B-cell differentiation in a patient with a novel LRBA mutation

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Protocol publication

[…] The Illumina paired-end genomic DNA sample preparation kit (PE-102-1001, Illumina, San Diego, CA, USA) was used for preparing the libraries including end repair, A-tailing and ligation of the Illumina adaptors. Each sample was prepared with an index using the Illumina multiplexing sample preparation oligonucleotide kit (PE-400-1001, Illumina) and then pooled in batches of six in equimolar amounts prior to exome enrichment. The Illumina TruSeq exome kit (FC-121-1008, Illumina) was used to capture the human exome for each sample pool. Each 6-plex exome enriched library was sequenced in two lanes of an Illumina HiSeq 2000 with version 2 chemistry as 100 bp paired-end reads.Sequence reads were mapped to the GRCh37 assembly of the reference human genome using the default parameters of the Burrows–Wheeler Aligner (bio-bwa.sourceforge.net). Untrimmed reads were aligned allowing a maximum of two sequence mismatches and reads with multiple mappings to the reference genome were discarded along with PCR duplicates. Sequence variants were identified with SAMtools (samtools.sourceforge.net) and annotated using Annovar (http://annovar.openbioinformatics.org). […]

Pipeline specifications

Software tools BWA, SAMtools, ANNOVAR
Application WES analysis
Organisms Homo sapiens