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Pear

Pear is a tool to merge paired-end sequencing reads, prior to downstream tasks such as assembly.

Get data

Input: paired-end reads.

  • We will use a set of Illumina MiSeq reads from the bacteria Staphylococcus aureus.

Go to your Galaxy server.

  • In the tool panel, go to Get Data: Upload File
  • Select Paste/Fetch data
  • In the box, paste in:

ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR171/008/ERR1712338/ERR1712338_2.fastq.gz ftp://ftp.sra.ebi.ac.uk/vol1/fastq/ERR171/008/ERR1712338/ERR1712338_1.fastq.gz

  • Click Start and then Close.
  • These two files will upload to your current Galaxy history.
  • Using the pencil icon, change the filetype to “fastqsanger”, and shorten the name of the file.

files

Run Pear

In the tool panel, go to NGS Analysis: NGS QC and manipulation: Pear

  • Dataset type: Paired-end
  • Name of file that contains the forward paired-end reads: ERR1712338_1.fastq
  • Name of file that contains the reverse paired-end reads: ERR1712338_2.fastq
  • Leave other settings as per defaults, except:
  • Maximal proportion of uncalled bases in a read: 0.01
    • omits reads if >1% of the reads is missing (N)
  • Output files: Select all

Your tool interface should look like this:

pear interface

  • Click Execute

Results

There are four output files.

  • Assembled reads: merged paired-end reads.
  • Unassembled forward reads and Unassembled reverse reads: remaining, unmerged reads.
  • Discarded reads: Did not meet quality specified

In this case, most of the reads have been merged (~360MB); 90MB are unmerged, and 350 sequences have been discarded.

Next

Run Trimmomatic to trim sequences before assembling.

Pear paper

Pear software