RNASeq023---HSCC-1-month-chronic-withdrawal

##Read Counts and Normalization of RNAS-Seq on HS Collaborative Cross (1-month EtOH chronic withdrawal experiment)

###The analysis related files are distributed among 3 directories:

• data: input data and intermediate files
  • data/Alignment_mm10 --> bam and bai files
  • data/Normalization --> all input files needed to generate normalized read counts
• scripts: all scripts used for analysis
• analysis: final analysis results files

Alignment:

####Since Sample file numbers had some breaks in sequence, I used 3 scripts to run the alignment simultaneously for different batches of samples.

In ACC local server:

nohup ./run_star_alignment_1.sh > ./run_star_alignment_1.log 2>&1 </dev/null &
nohup ./run_star_alignment_1b.sh > ./run_star_alignment_1b.log 2>&1 </dev/null &
nohup ./sort_and_index_1b.sh > ./sort_and_index_1b.log 2>&1 </dev/null &

In exacloud:

condor-submit align.sub

align.sub:

     executable              = run_star_alignment_2.sh
     log                     = submit_run_star_alignment.log
     output                  = submit_run_star_alignment.out
     error                   = submit_run_star_alignment.err
     request_cpus            = 20
     request_memory          = 64 GB
     notification            =Complete
     notify_user             [email protected]
     queue

Counts:

> nohup ../../scripts/run_bedtools_coverage.sh > ./run_bedtools_coverage.log 2>&1 </dev/null &
> mv RNASeq023_mm10_coverage_splitoption.txt ../../analysis/
> ls -d Sample_RNA150217RH_* > samples1.txt
> cd ../150423_D00735_0035_BC7D48ACXX/
> ls -d Sample_RNA150217RH_* >> ../150423_D00735_0034_AC791EACXX/samples1.txt
> cd ../150423_D00735_0034_AC791EACXX/

After removing unnecessary substrings from sample names and tab separating core id from lab id:

> mv samples1.txt sample_key.txt
> mv sample_key.txt ../data/
> cd ../data/
> sort -n sample_key.txt > sample_key_sorted
> mv sample_key_sorted sample_key_sorted.txt
> cd Alignment_mm10/
> ls Alignment_mm10/*sorted*bam > bam_files_counts_header_order.txt

Removed unnecessary substrings from sample names and tab separating core id from lab id

Normalization:

• used --> scripts/selectNormalizeGeneExonCounts_RNASeq023.R