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  • lootpiz
    Junior Member
    • May 2012
    • 1

    #1

    Produce different Tophat results with and without -z0 option

    Aligned paired-end RNA-seq using Tophat ver. 1.4.0.
    Used -z0 option for Tophat but it produced a different bam result.

    input fastq file size : 4.1G * 2 for paired-end
    ==================
    output 1. without -z0 option
    Code:
    $ tophat -r 94 -p 8 -G genes.gtf genome s_1_1.fastq s_1_2.fastq
    1.3G : accepted_hits.bam
    88K : deletions.bed
    50K : insertions.bed
    3.9M : junctions.bed
    70 : left_kept_reads.info
    4.0K : logs
    70 : right_kept_reads.info
    93M : unmapped_left.fq.z
    113M : unmapped_right.fq.z
    ==================
    output 2. with -z0 option
    Code:
    $ tophat -z0 -r 94 -p 8 -G genes.gtf genome s_1_1.fastq s_1_2.fastq
    523M : accepted_hits.bam
    55K : deletions.bed
    31K : insertions.bed
    2.6M : junctions.bed
    70 : left_kept_reads.info
    4.0K : logs
    70 : right_kept_reads.info
    2.7G : unmapped_left.fq
    2.5G : unmapped_right.fq
    ==================

    Then, I used samtools flagstat to look at bam file stats.
    The first bam file(without -z0 option) reported:
    Code:
    $ samtools flagstat accepted_hits.bam
    
    57149297 + 0 in total (QC-passed reads + QC-failed reads)
    0 + 0 duplicates
    57149297 + 0 mapped (100.00%:-nan%)
    57149297 + 0 paired in sequencing
    28845685 + 0 read1
    28303612 + 0 read2
    42295726 + 0 properly paired (74.01%:-nan%)
    54359924 + 0 with itself and mate mapped
    2789373 + 0 singletons (4.88%:-nan%)
    0 + 0 with mate mapped to a different chr
    0 + 0 with mate mapped to a different chr (mapQ>=5)
    The second bam file(with -z0 option) reported:
    Code:
    $ samtools flagstat accepted_hits.bam 
    
    20898015 + 0 in total (QC-passed reads + QC-failed reads)
    0 + 0 duplicates
    20898015 + 0 mapped (100.00%:-nan%)
    20898015 + 0 paired in sequencing
    10912428 + 0 read1
    9985587 + 0 read2
    3175304 + 0 properly paired (15.19%:-nan%)
    13747868 + 0 with itself and mate mapped
    7150147 + 0 singletons (34.21%:-nan%)
    0 + 0 with mate mapped to a different chr
    0 + 0 with mate mapped to a different chr (mapQ>=5)
    Does Tophat use different algorithm depend on -z0 option?
    How come Tophat produces different bam files?

    Thanks in advance!
    #####################
    # QUE LA FORCE SOIT AVEC TOI #
    #####################

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