Hi,
I'm trying to count reads mapping to sense strand. I have doubts which counts file I should chose from this pipeline. I think is "plate_R.counts" because has more reads counted in total. Am I right?
Library creation kit -> E7420S NEBNext® Ultra™ Directional RNA Library Prep Kit for Illumina®
I would also appreciate a nice tutorial to understand Illumina paired-end library preparation, alignment, counting...
Thanks!
P.S I read previous post asking similar questions but still I have doubts!
https://www.biostars.org/p/92935/
#################################################
#BWA HT.seq bacterial paired-end RNA-seq pipeline
#################################################
# Get the genome file from the command line
genome_file=$1
# Get the fastq file from the command line
fastq_file_R1=$2
# Get the fastq file from the command line
fastq_file_R2=$3
#get gff
GFF=$6
#BWA index (default settings)
bwa index $genome_file
#BWA align
bwa mem -t 8 $genome_file $fastq_file_R1 $fastq_file_R2 | gzip -3 > P_S1_L001_aln-pe.sam.gz
#Flagstat
#Convert .sam to .bam to input to Flagstat
samtools view -b -S -o P_S1_L001_aln-pe.bam P_S1_L001_aln-pe.sam.gz
samtools flagstat P_S1_L001_aln-pe.bam
#Count reads mapped with htseq-count
samtools sort -n P_S1_L001_aln-pe.bam plate.sorted
python -m HTSeq.scripts.count -m intersection-nonempty -f bam -a 10 -t mRNA -i Parent -r name -s yes plate.sorted.bam $GFF | awk 'n>=5 { print a[n%5] } ...
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