Illumina MiSeq Results and Troubleshooting

The MiSeq produces various output files depending on the workflow requested by the user.

The majority of runs produce a FASTQ file that comprises the sequences produced by the run as well as subsequent quality scores of each base. The cumulative size of FASTQ files for a typical MiSeq run ranges from 1-4 Gb of data.

More information on the MiSeq applications can be found at http://www.illumina.com/systems/miseq/applications.ilmn

 

Technical support will be provided on an as-needed basis for construction of requested libraries and sequencing of Illumina libraries at a predetermined charge.

Typical issues and resolutions include:

 

Problem

Cause

Solution

Library Over Clustering

 

Under quantification of library prior to sequencing

 

Re-quantify library by Bioanalyzer, fluorometric dsDNA assay (PicoGreen or Qubit), or Illumina recommended Q-PCR assay. Once re-quantified, sequence again after diluting to a lesser degree based off of new quantification results. 

Library Under Clustering

 

Over quantification of library prior to sequencing

Re-quantify library by Bioanalyzer, fluorometric dsDNA assay (PicoGreen or Qubit), or Illumina recommended Q-PCR assay. Once re-quantified, sequence again after diluting to a greater degree based off of new quantification results. 

Low Passing Filter Percentage

 

Homogeneous library

Add a Heterogeneous library to your sequencing run. Typically libraries that are more heterogeneous sequence better on the MiSeq. Thus by adding a heterogeneous library to a sequencing run, the quality of data produced increases. This can be achieved by increasing the percentage of PhiX Control Library added to a run to a maximum of 50%.

 

 

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