Fast-GBS Demultiplexing with Sabre for PAIRED-END READS

Question

[csjlee08]

Hello - I am trying to analyze my sequencing data and am having trouble with the demultiplex step of Fast-GBS, which utilizes Sabre. The problem appears to be with the demultiplex step. I used paired-end sequencing method so I have two barcode files. Because of this I have named them barcodes_FLOW1_1 and barcodes_FLOW1_2 and set the lanes to two lanes, although the MiSeq only has one flow cell and one lane. The sequence data is names FLOW1_1.fq.gz and FLOW1_2.fq.gz. I have used just the 5’-3’ sequence as there is no other place to put the second adapter sequence. I have checked the names of the files and all appears to be fine, I removed the .txt from the file name as instructed.

The error shown on terminal is:

Demultiplex with Sabre
./fastgbs.sh: line 213: parallel: command not found
    !!! There is a problem with the demultiplex step with sabre for 
    !!! Check the names given to the sequences and barcode files

Here is the parameters I have set:

; FLOWCELL INFORMATION 
FLOWCELL=FLOW1 

; LANES INFORMATION 
FLOW1_LANES=1 2

; SEQUENCING TECHNOLOGY: ILLUMINA IONTORRENT 
TECHNOLOGY=ILLUMINA 

; SEQUENCE TYPE (PAIRED_END OR SINGLE-END): PE SE 
SEQTYP=PE 

; NAME OF THE REFERENCE GENOME FILES 
REFGEN=adzukireferencegenome.fasta 

; SEQUENCE USED FOR ADAPTOR RECOGNITION (THE COMPLETE ADAPTOR SEQUENCE IS 
; AGATCGGAAGAGCGGG) 
ADAP=CACGACGCTCTTCCGATCT

; MINIMUM READ LENGTH TO KEEP 
READLEN=50 

; NUMBER OF PARALLEL PROCESS FOR BWA 
BWAPAR=2 

; NUMBER OF THREAD FOR EACH BWA PROCESS 
BWATHR=4 

; MAXIMUM NUMBER OF CORES THAT COULD BE ALLOWED BY YOUR COMPUTER FOR THIS ANALYSIS 
; SHOULD BE EQUAL TO: BWAPAR X BWATHR 
NBCOR=8 

; NAME OF FILE CONTAINING LOG 
LOGFILE=logfile_fastgbs.log 

; NAME OF FILE CONTAINING LIST OF S1.BAM FILE FOR PLATYPUS ANALYSIS  
BAMLIST=List_bam 

; FOR PLATYPUS: Minimum number of supporting reads required before a 
; variant candidate will be considered (equivalent to minDP in vcftools) 
MINREADS=2 

; PLATYPUS LOGFILE 
LOGPLAT=FastGBS_platypus_log.txt 

; PLATYPUS OUTPUT VCF FILE 
OUTPLAT=FastGBS_platypus 

; VCFTOOLS filtering: --max-missing 0.2 
MAXMIS=0.2

Answer 1

[Pierre Lindenbaum]

parallel: command not found

parallel is not installed or it is not in your PATH....

[csjlee08]

I installed parallel, seems to have gotten rid of that error code, however I checked the shell script and it is written for single end not paired end reads so I have changed the parameters to match the paired end reads but no luck. I get a new error:

/bin/bash sabre: command not found
/bin/bash sabre: command not found

printf "\nDemultiplex with sabre\n" | tee -a "${logfile}"
Step=$(grep "SABRE" checkpoint_${1})
if [ "${Step}" != "SABRE" ]
    then
        cd data
        parallel -j "${nbcor}" sabre pe -f FLOW1_1.fq.gz -r FLOW1_2.fq.gz -b barcodes_FLOW1.txt -u unknown_barcode1.fastq -w unknown_barcode1.fastq ::: $(ls -1 *.fq.gz | sed 's/.fq.gz//')
        if [ $? -ne 0 ]
            then 
                printf "\t!!! There is a problem in the demultiplex step with sabre for ${f} ${l}\n" | tee -a ../"${logfile}"
                printf "\t!!! Check the names given to the sequences and barcode files\n" | tee -a ../"${logfile}"
                exit 1

[Pierre Lindenbaum]

it's the very same kind of error....