Question: Both Paired And Unpaired Read Data Input For Subread Rna Sequencing Read Alignment
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gravatar for alpha2zee
5.2 years ago by
alpha2zee100
alpha2zee100 wrote:

Does anyone know if the Subread RNA seq. aligner software accepts both paired and unpaired read data-files for the same sample, the way Tophat does? The Subread manual doesn't say anything on this.

In my tests, Subread doesn't accept comma-separated paired and unpaired file-names (Tophat's usage), but it seems to accept space-separated file-names, and generates BAM files.

However, I am not sure if it actually reads the unpaired data. As per ngsutils bamutils stat.py, the BAM files produced with inputs that had or did not have the unpaired data seem to be exactly identical (15563374 and 14117543, and 15563374 and 14117543 total and mapped reads, respectively).

My samples have both paired and unpaired data because I use trimmomatic to process the initial all-paired data to clean/filter reads to remove adapter sequences, trim reads of poor-quality trailing bases, etc.

# comma separation of file-names in case of Tophat (tophat)
... -o output index paired_1.fastq,unpaired_1.fastq paired_2.fastq,unpaired_2.fastq

# comma separation of file-names fails for Subread (subjunc or subread-align)
... -i index -r paired_1.fastq,unpaired_1.fastq -R paired_2.fastq,unpaired_2.fastq -o output

# but space separation of file-names seems to work
... -i index -r paired_1.fastq unpaired_1.fastq -R paired_2.fastq unpaired_2.fastq -o output

# though output.bam is almost identical to one produced without the unpaired data
... -i index -r paired_1.fastq -R paired_2.fastq -o output
alignment rna-seq • 2.9k views
ADD COMMENTlink modified 5.2 years ago • written 5.2 years ago by alpha2zee100
1
gravatar for alpha2zee
5.2 years ago by
alpha2zee100
alpha2zee100 wrote:

Subread, as of version 1.4.3-p1, does not accept both paired and unpaired data for the same sample... see https://groups.google.com/forum/#!topic/subread/3tkBGjz6zVI

ADD COMMENTlink written 5.2 years ago by alpha2zee100
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