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Consumable FAQ for the Illumina 5 Base Whole Genome Sequencing (WGS) and Enrichment kits

What are the kit configurations for the Whole Genome Sequencing (WGS) and Enrichment kits?

WGS

  • 24 sample kit

  • Indexes required, but sold separately

Enrichment

  • 24 sample enrichment kit

  • Indexes required, but sold separately

What are the key/uncommon requirements for user-supplied consumables and equipment?

WGS or Enrichment kits:

  • [gDNA] Covaris LE220 Plus, or other sonication equipment + tubes

  • [cfDNA] Agilent Fragment Analyzer or Agilent TapeStation + reagents for input quantification

  • Hard-shell PCR plates (250 ul min volume)

  • DynaMag Magnet (+/- skirted)

  • Qubit + Qubit dsDNA Broad Range kit

  • High-Speed Thermal Mixer, or equivalent instrument that can shake up to 2200 rpm

  • Note: No additional equipment or consumables are required for the Enrichment workflow compared to the WGS kit, outside of purchasing an enrichment panel of oligos. Hybridization is performed on a thermal cycler.

  • Consult the library prep Product Documentation for a full list of required equipment and consumables.

Which enzyme is the IME conversion enzyme?

  • The details of the enzyme are proprietary to Illumina.

Can strip tubes be used instead of 96 well plates?

  • No, Illumina does not recommend using strip tubes. They do not have the same thermal conductance as 96 well plates, which could make thermal cycling less robust. This could lead to poor methylation conversion, for instance. In addition, all of the shaking steps have been optimized for plates, not tubes.

What is the supported number of freeze thaws of Illumina reagents?

  • 3 freeze/thaws.

Are any reagents very sensitive in this protocol? Eg, to freeze-thaw, precipitates, etc.?

  • The Illumina Methylation Enzyme (IME, the conversion enzyme) is sensitive to ethanol contamination and bead carryover. It is critical to make sure there is no residual ethanol or beads in the sample before the conversion step.

  • IME can be staged on ice for >30 minutes when running the protocol. But it should be stored long term at the recommended temp of -20°C, otherwise its conversion activity will be reduced.

Can the volume of MRR be modified?

  • No, modifications to the MRR volume can have negative effect on methylation conversion and therefore should not be altered.

For any feedback or questions regarding this article (Illumina Knowledge Article #9940), contact Illumina Technical Support [email protected].

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