How to set up a PhiX validation run on the NextSeq 500/550 in manual mode

A PhiX validation run confirms proper hardware and software performance of the instrument. The Illumina PhiX control library is a well-balanced genome with relatively equal representation of A, C, G, and T nucleotides. PhiX lacks an index and is not an appropriate tool for assessing Index Read performance. Below are instructions for setting up a PhiX validation run for the NextSeq 500/550 system in Manual mode. Manual mode allows you to setup the run without using Local Run Manager or BaseSpace.
  1. 1.
    Open the NextSeq 500/550 Control Software (NCS).
  2. 2.
    Select Experiment.
  3. 3.
    Select Sequence.
  4. 4.
    Select Manual.
  5. 5.
    Edit the run parameters in Run Setup; an example paired-end run is shown below.
Note: For the run, set both Index 1 and Index 2 to zero.
  1. 7.
    Follow the prompts on the screen to load the flow cell.
  2. 8.
    Follow the prompts on the screen to discard spent reagents, reload the spent reagents tray, and load a new buffer cartridge.
  3. 9.
    Load the reagent cartridge.
  4. 10.
    Review the run configuration.
  5. 11.
    Next, perform the instrument self-check, after which the PhiX validation run begins sequencing.
For more information about preparing PhiX for the NextSeq 500/550 sequencing platform, see the following documents:
For any feedback or questions regarding this article (Illumina Knowledge Article #1532), contact Illumina Technical Support [email protected].
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