How to set up a PhiX validation run on MiSeq i100 Series
Background
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, T, G, and C nucleotides. PhiX lacks an index and is not an appropriate tool for assessing Index Read or DRAGEN Secondary Analysis performance.
Below are instructions for setting up a PhiX validation run on MiSeq i100 series sequencing platforms. The optimal loading concentration for a PhiX validation run on the MiSeq i100 series is 120 pM.
Plan a Sequencing Run
Cloud run planning:
To plan run in the Cloud using BaseSpace Sequence Hub, use the Run Planning tool in BaseSpace Sequence Hub to set up a sequencing run. For more information on BaseSpace Sequence Hub, refer to the BaseSpace Sequence Hub support site page.
Local run planning:
Select the menu icon in the upper-left corner.
Select Runs.
On the Planned tab, select Create run.
Enter a run name to identify the run.
Enter the number of cycles performed in each read:
For the 100 cycle kit, use 51 cycles for Read 1 and Read 2
For the 300 cycle kit, use 151 cycles for Read 1 and Read 2
For the 600 cycle kit, use 301 cycles for Read 1 and Read 2.
Set number of cycles for Index 1 and Index 2 to 0 (zero) for all kits. Do not leave these fields blank.
Select Next.
Select the analysis application, typically DRAGEN BCL Convert.
Select the library prep and index adapter kits, typically Not Specified.
Select Next.
Input the Sample ID, typically PhiX.
Select the FASTQ file compression format, typically gzip.
Select Next.
Select Save as Planned.
Start a Sequencing Run
Select Start.
Sign in with local or cloud account.
Select the PhiX run configured in the steps above from the Planned Runs list.
Select Review and then review your run information.
Select Load consumables.
Place the new dry cartridge in the dry cartridge tray. Gently push the dry cartridge until it touches the back of the tray, so it is firmly secured.
Select Next.
Remove the plastic cap and load the wet cartridge.
Select Close.
Select Verify Run.
Confirm the run and consumables are correct, and then select Start run.
For any feedback or questions regarding this article (Illumina Knowledge Article #9320), contact Illumina Technical Support techsupport@illumina.com.
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