documentation

After successful installation of the package use

$ htseq-clip -h

for a brief description of the functions available in htseq-clip. The available functions can be categorized into 4 different classes given below.

Prepare annotation

annotation

Flattens a given annotation file in GFF format to BED6 format

Arguments

• -g/--gff GFF formatted annotation file, supports .gz files
• -u/--geneid Gene id attribute in GFF file (default: gene_id)
• -n/--genename Gene name attribute in GFF file (default: gene_name)
• -t/--genetype Gene type attribute in GFF file (default: gene_type)
• --splitExons This flag splits exons into components such as 5’ UTR, CDS and 3’ UTR
• --unsorted Use this flag if the GFF file is unsorted
• -o/--output Output file name. If the file name is given with .gz suffix, it is gzipped. If no file name is given, output is print to console

Note

The default values for --geneid, --genename and --genetype arguments follow gencode GFF format

Usage

$ htseq-clip annotation -h

createSlidingWindows

Create sliding windows from the flattened annotation file

Arguments

• -i/--input Flattened annoation file, see annotation
• -w/--windowSize Window size in number of base pairs for the sliding window (default: 50)
• -s/--windowStep Window step size for sliding window (default: 20)
• -o/--output Output file name. If the file name is given with .gz suffix, it is gzipped. If no file name is given, output is print to console

Usage

$ htseq-clip createSlidingWindows -h

mapToId

Extract “name” column from the annotation file and map the entries to unique id and print out in tab separated format

Arguments

• -a/--annotation Flattened annotation file from annotation or sliding window file from createSlidingWindows
• -o/--output Output file name. If the file name is given with .gz suffix, it is gzipped. If no file name is given, output is print to console

Usage

$ htseq-clip mapToId -h

Extract crosslink sites

extract

Extract crosslink sites, insertions or deletions

Arguments

• -i/--input Input .bam file. Input bam file must be co-ordinate sorted and indexed
• -e/--mate for paired end sequencing, select the read/mate to extract the crosslink sites from, accepted choices: 1, 2
  • 1 use the first mate in pair
  • 2 use the second mate in pair
• -s/--site Crosslink site choices, accepted choices: s, i, d, m, e (default: e)
  • s startsite,
  • i insertion site
  • d deletion site
  • m middle site
  • e end site
• -g/--offset Number of nucleotides to offset for crosslink sites (default: 0)
• --ignore Use this flag to ignore crosslink sites outside of genome annotations
• -q/--minAlignmentQuality Minimum alignment quality (default: 10)
• -m/--minReadLength Minimum read length (default: 0)
• -x/--maxReadLength Maximum read length (default: 500)
• -l/--maxReadInterval Maximum read interval length (default: 10000)
• --primary Use this flag consider only primary alignments of multimapped reads
• -c/--cores Number of cores to use for alignment parsing (default: 5)
• -t/--tmp Path to create and store temp files (default behavior: use parent folder from “–output” parameter)
• -o/--output Output file name. If the file name is given with .gz suffix, it is gzipped. If no file name is given, output is print to console

Usage

$ htseq-clip extract -h

Note

To extract 1``st offset position of second mate (``2) start site (s) in eCLIP, use: --mate 2 --site s --offset -1

Count crosslink sites

count

Counts the number of crosslink/deletion/insertion sites

Arguments

• -i/--input Extracted crosslink sites, see extract
• -a/--ann Flattened annotation file, see annotation OR sliding windows file, see createSlidingWindows
• --unstranded crosslink site counting is strand specific by default. Use this flag for non strand specific crosslink site counting
• -o/--output Output file name. If the file name is given with .gz suffix, it is gzipped. If no file name is given, output is print to console

Usage

$ htseq-clip count -h

Helper functions

createMatrix

Create R friendly output matrix file from count function output files

Arguments

• -i/--inputFolder Folder name with output files from count function, see count
• -b/--prefix Use files only with this given file name prefix (default: None)
• -e/--postfix Use files only with this given file name postfix (default: None)
• -o/--output Output file name. If the file name is given with .gz suffix, it is gzipped. If no file name is given, output is print to console

Warning

either --prefix or --postfix argument must be given

Usage

$ htseq-clip createMatrix -h

createMaxCountMatrix

Create R friendly output matrix file from crosslink_count_position_max column in count function output files. This file can be used to filter down the output file from createMatrix function during downstream statistical analysis.

Arguments

• -i/--inputFolder Folder name with output files from count function, see count
• -b/--prefix Use files only with this given file name prefix (default: None)
• -e/--postfix Use files only with this given file name postfix (default: None)
• -o/--output Output file name. If the file name is given with .gz suffix, it is gzipped. If no file name is given, output is print to console

Warning

either --prefix or --postfix argument must be given

Usage

$ htseq-clip createMatrix -h