Computational protocol: Mutations in epigenetic regulators including SETD2 are gained during relapse in pediatric acute lymphoblastic leukemia

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Protocol publication

[…] Fastq files were aligned to hg19 version of the human genome with BWA. SNP, indel and copy number variant calling was performed with samtools mpileup and Varscan 2.2.3 somatic option. Annovar was used to annotate variants. Relapsed and diagnostic variants were linked and variants were filtered for >=15% allele frequency and 20x coverage in either tumor sample, <5% and 10x coverage in the remission sample, bidirectional variant reads, minimal strand bias, and minimal local noise. Variants were visualized in the Integrated Genome Viewer to eliminate variants with clustered read position and variants caused by indel artifacts. Sanger sequencing was used to validate a sample of somatic variants found by next generation sequencing to validate filtering criteria. Every filtered somatic variant position was then forced called on every sample to examine the baseline noisiness of each position. Those with mean baselines >1% allele frequency in remission samples were excluded. Variants were also filtered to only those positions within 50 base pairs of those included in the custom capture baitset. Variants were considered somatic if they were present in less than 5% allele frequency in the paired remission sample (to account for high minimal residual disease). […]

Pipeline specifications

Software tools BWA, SAMtools, VarScan, ANNOVAR, IGV
Application Genome data visualization
Organisms Homo sapiens