[[TOC()]]

= Coverage Analysis Pipeline =
TODO. Suggested parties to take this up: Antoine van Kampen, Barbera van Schaik, Silvia D Olabarriaga, Mark Santcroos, AMC

= Workflows =

== Create grid directory and change permissions ==

[[Image(CreateGridDirectory.png)]]
   * Creates a directory on the LFC
   * Changes the permissions such that it is in-accessible to the group and others

== Create a BWA index on database ==

[[Image(bwaIndexDatabase.png, 50%)]]

Gunzip fasta file. Build BWA index. Tar-gzip the results.

== Split fastq file ==

[[Image(splitFastq.png, 50%)]]

Splits a large fastq file (gzipped) into several smaller files with the unix command 'split'. The results are uploaded to the directory that is specified in 'gridOutputDir'

== Alignment with BWA on each split file ==

[[Image(BWAparam.png, 50%)]]

Runs BWA with adjustable parameter settings.
   * Matches sequence reads to a reference database
   * Convert sai to sam
   * Convert sam to bam
   * Sort bam file
   * Index sorted bam file
   * Tar-gzip all results. Also the intermediate files

== Merge bam files ==

[[Image(MergeIndexSNPcall.png, 50%)]]

   * Downloads all bai, bam, sam and tar.gz files from the gridInputDirectory
   * Gunzip tar the tar.gz files if they are present
   * Gunzip the reference file (fasta format)
   * Merge all _sorted.bam files
   * Build index on this merged file
   * Call SNPs and make selection. Output in pileup format.
   * Convert pileup format to bed format

== SNP calling with varscan, determine coverage ==

[[Image(Coverage_Varscan_BaseCoverage.png)]]

   * Creates a pileup file (with samtools pileup -f) Sends the output to Varscan. Calls SNPs, indels and copy number variations.
   * Calculates coverage per 50kbp
   * Calculates coverage per base