Last updated: 2018-06-13

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    File Version Author Date Message
    Rmd 642faf0 Briana Mittleman 2018-06-13 start PAS enrichment analysis

I am going to use this analysis to look for enrichment of my 3’ seq reads at annoated PAS sites. This is similar to the analysis I ran for the net-seq https://brimittleman.github.io/Net-seq/use_deeptools.html.

Running Deep Tools:

Step 1: Create bigwig coverage files with bamcoverage

  • bamCoverage -b reads.bam -o coverage.bw

Step 2: computeMatrix

I will need my normalized bigwig reads and the bed interval file (in my case PAS clusters)

ex: computeMatrix scale-regions -S -R -b 1000 -a 1000 -out

–skipZeros (option- not included in first try)

Step 3: Plot heatmap

required –matrixFile, -m (from the compute matrix), -out (file name to save image.png)

–sortRegions descending

–plotTitle, -T

#!/bin/bash


#SBATCH --job-name=deeptools_pas
#SBATCH --time=8:00:00
#SBATCH --partition=broadwl
#SBATCH --mem=40G
#SBATCH --tasks-per-node=4 
#SBATCH --mail-type=END
#SBATCH --output=deeptool_pas_sbatch.out
#SBATCH --error=deeptools_pas_sbatch.err

module load Anaconda3

source activate three-prime-env

sample=$1
describer=$(echo ${sample} | sed -e 's/.*\YL-SP-//' | sed -e "s/-sort.bam$//")


bamCoverage -b $1
 -o /project2/gilad/briana/threeprimeseq/output/deeptools/${describer}.bw

computeMatrix reference-point -S project2/gilad/briana/threeprimeseq/output/deeptools/${describer}.bw  -R /project2/gilad/briana/apa_sites/rnaseq_LCL/clusters_fullAnno.bed  -b 500 -a 500 -out /project2/gilad/briana/threeprimeseq/output/deeptools/${describer}.PAS.gz

plotHeatmap --sortRegions descend --refPointLabel "PAS"  -m /project2/gilad/briana/threeprimeseq/output/deeptools/${describer}.PAS.gz  -out /project2/gilad/briana/threeprimeseq/output/deeptools/${describer}.PAS.gz.png

I am running this on YL-SP-18486-N_S10_R1_001-sort.bam to try it first.

Session information

sessionInfo()
R version 3.4.2 (2017-09-28)
Platform: x86_64-apple-darwin15.6.0 (64-bit)
Running under: macOS Sierra 10.12.6

Matrix products: default
BLAS: /Library/Frameworks/R.framework/Versions/3.4/Resources/lib/libRblas.0.dylib
LAPACK: /Library/Frameworks/R.framework/Versions/3.4/Resources/lib/libRlapack.dylib

locale:
[1] en_US.UTF-8/en_US.UTF-8/en_US.UTF-8/C/en_US.UTF-8/en_US.UTF-8

attached base packages:
[1] stats     graphics  grDevices utils     datasets  methods   base     

loaded via a namespace (and not attached):
 [1] workflowr_1.0.1   Rcpp_0.12.17      digest_0.6.15    
 [4] rprojroot_1.3-2   R.methodsS3_1.7.1 backports_1.1.2  
 [7] git2r_0.21.0      magrittr_1.5      evaluate_0.10.1  
[10] stringi_1.2.2     whisker_0.3-2     R.oo_1.22.0      
[13] R.utils_2.6.0     rmarkdown_1.8.5   tools_3.4.2      
[16] stringr_1.3.1     yaml_2.1.19       compiler_3.4.2   
[19] htmltools_0.3.6   knitr_1.18

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