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# Standard operating procedure - Alignment, variant calling, and annotation of trio whole exome samples at the Edinburgh Parallel Computing Centre

This SOP applies to batches of family/trio samples where trio whole exome sequencing has been performed by Edinburgh Genomics (EdGE). It assumes that data has been successfully transferred from EdGE to the Edinburgh Parallel Computing Centre (EPCC) (see SOP: Transfer of whole exome sequencing samples from Edinburgh Genomics to Edinburgh Parallel Computing Centre). Scripts are version controlled on the University of Edinburgh gitlab server gitlab.ecdf.ed.ac.uk/igmmbioinformatics/trio-whole-exome. Request access by e-mail: alison.meynert@igmm.ed.ac.uk.

## Definitions

In this document, N is the total number of samples in the project, and X is the number of families.

Text in angle brackets, e.g. <project> indicates variable parameters. A variable parameter such as <family1-X> indicates that there are X instances of the parameter, each with their own unique value.

## Software and data requirements

The analysis is run with the bcbio pipeline (version 1.1.15) located at /home/u035/project/software/bcbio. All genome reference and annotation data resources are contained within the genomes/Hsapiens/hg38 subfolder.

The TWIST target BED file is at: /home/u035/project/resources/Twist_Exome_RefSeq_targets_hg38.plus15bp.bed

To generate the target BED file, first copy the file Twist_Exome_RefSeq_targets_hg38.bed from NHS Clinical Genetics Services to /home/u035/project/resources on ultra, then pad it by 15bp each side.

```
cd /home/u035/project/resources

../software/bcbio/tools/bin/bedtools slop -g \
  ../software/bcbio/genomes/Hsapiens/hg38/seq/hg38.fa.fai \
  -i Twist_Exome_RefSeq_targets_hg38.bed -b 15 | \
  ../software/bcbio/tools/bin/bedtools merge > \
  Twist_Exome_RefSeq_targets_hg38.plus15bp.bed
```