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Control (GDMC and SFMC) FAQ for the Illumina 5 Base Whole Genome Sequencing and Enrichment kits

What controls are needed in the prep? What is the GDMC (Genomic DNA Methylation Control) and SFMC (Short Fragment Methylation Control)? Why are they mixed with the sample? How do they work in the assay?

  • GDMC is a combination of pUC19 (a fully methylated genome) and Lambda (a fully unmethylated genome). It is mixed with the sample prior to shearing to help judge the conversion success of the sample.

  • SFMC is the same pUC19 and Lamba genomes; however, it is pre-sheared to mimic cfDNA and is spiked into cfDNA samples prior to starting the workflow.

Is it correct that the only difference between SFMC and GDMC is the fragment size (ie, SFMC is sheared to match cfDNA fragment size and gDNA is high molecular weight control DNA)?

  • This is correct.

Are the controls added to each sample (eg, to all 96 samples on a plate) or are they a separate sample (eg, 2 of 96 samples on the plate are controls?) Does this change for the Enrichment workflow?

  • Each gDNA sample is combined with 2.5 µl of GDMC (Genomic DNA Methylation Control).

  • Each cfDNA sample is combined with 2.5 µl SFMC (Short Fragment Methylation Control).

  • The spike-in controls will be spiked in with the DNA samples and are intended to be library-prepared and sequenced along with the library. The customer will not need to sequence them separately.

  • This is the same for both the WGS and the Enrichment kit workflows; however, for the Enrichment workflow, to detect GDMC or SFMC, the Methylation Control Panel of oligos must be also purchased and used in the hybridization. For more details, see “Methylation Control Panel” section below.

Is an unconverted version of each sample needed in addition to the enzyme-converted sample for analysis purposes?

  • No, this is not required for the analysis.

Is it known how alternative controls would perform?

  • Alternate controls are not recommended. Current DRAGEN software is designed to use the Illumina-provided controls only.

For any feedback or questions regarding this article (Illumina Knowledge Article #9943), contact Illumina Technical Support [email protected].

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