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Bowtie giving opposite results
Hi all,
I am using the bowtie program to run alignments on a set of mate pair data (2 fastq files) and have run into a problem. What I have found is that if I run bowtie with the regular fastq files 'as is' with the --rf (MP orientation) option, then the results I get are "backwards", in the sense that reads with large insert sizes of 1000+ are being given bitFlags that indicate paired end orientation, and reads with smaller insert sizes of around 180 are being given mate pair orientation when they are clearly paired end. I also ran bowtie with the reverse complemented fastq files (accomplished this using a python script that reverse complemented the bases, and reversed the quality scores), and this time, bowtie2 gave bitFlags that made sense with the data, i.e. expected mate pair reads were being given bitFlag definitions of mate pair orientation. I am not sure if this is expected, and I should be giving bowtie the reverse complements of these fastq files, or if I have made a mistake in another option? I did specify '--rf' to indicate the data was MP, and gave -I and -X options.
Thanks to anyone who takes the time to read and respond to this post!
Riley
I am using the bowtie program to run alignments on a set of mate pair data (2 fastq files) and have run into a problem. What I have found is that if I run bowtie with the regular fastq files 'as is' with the --rf (MP orientation) option, then the results I get are "backwards", in the sense that reads with large insert sizes of 1000+ are being given bitFlags that indicate paired end orientation, and reads with smaller insert sizes of around 180 are being given mate pair orientation when they are clearly paired end. I also ran bowtie with the reverse complemented fastq files (accomplished this using a python script that reverse complemented the bases, and reversed the quality scores), and this time, bowtie2 gave bitFlags that made sense with the data, i.e. expected mate pair reads were being given bitFlag definitions of mate pair orientation. I am not sure if this is expected, and I should be giving bowtie the reverse complements of these fastq files, or if I have made a mistake in another option? I did specify '--rf' to indicate the data was MP, and gave -I and -X options.
/home/rcorbett/aligners/bowtie2/bowtie2-2.0.0-beta7/bowtie2 -p 8 --rf -I 2000 -X 2500 -x /home/rcorbett/aligners/bowtie2/bowtie2-2.0.0-beta7/hg19a -1 /projects/rcook/3_prj/bwa/30NE8AAXX_3_1_export_f1b6.fastq -2 /projects/rcook/3_prj/bwa/30NE8AAXX_3_2_export_f1b6.fastq -S /projects/rcook/3_prj/bowtie.align.output.rf.min2000.max2500.sam
Riley
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