2020 Analysis with bedtools
BED format
BED format is a simple 3 to 9 column format for location-oriented data.
See supported data formats for custom tracks for more information and examples.
Important rules in BED format
- The number of fields per line must be consistent throughout any single set of data in an annotation track.
- The first base in a chromosome is numbered 0
BED format practice 1
Q: Convert saccharomyces_cerevisiae_R64-1-1_20110208.gff into a 3 column bed file that includes 'gene' feature
BED format practice 2
Q: Convert saccharomyces_cerevisiae_R64-1-1_20110208.gff into a 4 column bed file that includes 'gene' feature (4th column has gene IDs)
BEDTools multicov
BEDTools is a great utility for comparing genomic features in BAM, BED, VCF, and GFF formats. The documentation is well written in great detail. Here, we will practice three commonly used sub-commands: multicov, merge, and intersect.
We are going to use it to count the number of reads that map to each gene in the genome. Load the module and check out the help for bedtools and the multicov specific command that we are going to use:
module load bedtools bedtools bedtools multicov
The multicov command takes a feature file (GFF/BED/VCF) and counts how many reads are in certain regions from many input files. By default it counts how many reads overlap the feature on either strand, but it can be made specific with the -s
option.
Note: Remember that the chromosome names in your gff file should match the way the chromosomes are named in the reference fasta file used in the mapping step. For example, if BAM file used for mapping contains chr1, chrX etc, the GFF file must also call the chromosomes as chr1, chrX and so on.
In order to use the bedtools command on our data, do the following:
bedtools multicov -bams yeast_chip_sort.bam yeast_chip2_sort.bam -bed sc_gene.bed > gene_counts.txt
Then take a peek at the data...
head gene_counts.txt
BEDTools merge
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BEDTools intersect
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Online tutorial
Overview and use cases
http://bedtools.readthedocs.org/en/latest/content/bedtools-suite.html
bedtools intersect
http://bedtools.readthedocs.org/en/latest/content/tools/intersect.html
bedtools multicov
http://bedtools.readthedocs.org/en/latest/content/tools/multicov.html
bedtools merge
http://bedtools.readthedocs.org/en/latest/content/tools/merge.html
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