My simple test for isoSeq3

from https://github.com/PacificBiosciences/IsoSeq3/blob/master/README.md

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step1 - CCS

ccs sub-soft from smrt link v5.1.0

ccs \
--force \
--minSnr 4 \
--minLength 200 \
--minPasses 1 \
--minPredictedAccuracy 0.8 \
--minZScore -999 \
--maxDropFraction 0.8 \
--numThreads 20 \
--reportFile ccs_report.txt \ #output a CCS summary file
--logLevel DEBUG \
--logFile ccs_log.txt \       #output a CCS process log file
--noPolish \
test.subreads.bam \           #input a original subreads.bam of sequel
ccs.bam                       #output CCS sequence file of BAM format

output files

ccs.bam ccs.bam.pbi ccs_log.txt ccs_report.txt

step2 - lima

easy to use from https://github.com/PacificBiosciences/IsoSeq3/releases/download/v0.7.2/isoseq3

 lima \
    ccs.bam \           #input CCS sequence file of step1
primers.fasta \     #input a file containing primers of 5p and 3p
demux.bam \         #output BAM#
--isoseq  \
--no-pbi  \
--dump-clips     \
--min-passes 1   \
--min-length 200 \
--chunk-size 20  \
--num-threads 20

NOTE:

   The true output name of demux.bam is demux.primer_5p--primer_3p.bam, which renamed by the name of primer

output files

    demux.json
    demux.lima.report
    demux.primer_5p--primer_3p.subreadset.xml
    demux.lima.clips
    demux.lima.summary

primers.fasta

    demux.lima.counts
    demux.primer_5p--primer_3p.bam

primers.fasta

    >primer_5p
    AAGCAGTGGTATCAACGCAGAGTACATGGGG

    >primer_3p
    AAGCAGTGGTATCAACGCAGAGTAC

---

step3 - cluster & polish (FAST and easy to use)

easy to use from https://github.com/PacificBiosciences/IsoSeq3/releases/download/v0.7.2/isoseq3

    isoseq3 cluster   \
    	demux.primer_5p--primer_3p.bam \  #input of step2
        unpolished.bam \                  #output
    	--verbose \
        --num-threads 20 \
        --logFile cluster.log

output files

    cluster.log
    unpolished.flnc.consensusreadset.xml
    unpolished.flnc.bam.pbi
    unpolished.flnc.bam
    unpolished.transcriptset.xml
    unpolished.bam.pbi

unpolished.bam

    unpolished.fasta
    unpolished.cluster

    isoseq3 polish \
    	unpolished.bam \      #input unpolished BAM file of cluster   
    	test.subreads.bam \   #input original subreads BAM file
    	polished.bam \        #output polished BAM
    	--verbose \
    	--num-threads 20 \
    	--logFile polish.log  #output plish LOG file

output files

    polished.lq.fastq.gz
    polished.hq.fastq.gz
    polished.lq.fasta.gz
    polished.hq.fasta.gz
    polished.transcriptset.xml
    polished.bam.pbi
    polished.bam
    polish.log