User's Manual

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Evaluate the enrichment of target DNA
After quantification of total DNA continue with measurement of target enrichment. An online tool for
calculating the DNA enrichment based on qPCR is available at samplix.com
To determine fold enrichment of target DNA, perform a qPCR using the qPCR QC primers, not overlapping
with the dPCR enrichment amplicon(s). See “Droplet PCR Reaction Design for Target Enrichment” (Chapter 2)
or the online primer design tool at samplix.com. If a specific region of DNA is required, place the validation
primer pair on the opposite side of this region but with maximum 5 kb distance from the dPCR primers. The
enrichment measure is indicative of enrichment and might differ from the enrichment measured by
sequencing.
To quantify the enrichment, perform a standard qPCR reaction using either the Samplix primer test PCR kit
(Cat. No. RE10200) or your own preferred qPCR reagents (Fig. 6.2).
Set up a qPCR for detection using the following DNA as a template:
• dMDA amplified sorted positive population, 1:10 dilution (enriched sample, red curve in Fig. 6.2)
• dMDA H
2
O, 1:10 dilution (negative control)
• Original sample input DNA in the same concentration as input in droplets. Run a standard curve of
original sample to calculate PCR efficiency (grey curves in Fig. 6.2).
Suggested controls:
• dMDA sheath fluid from flow cytometer, 1:10 dilution (contamination control)
• dMDA 1 pg non-sorted input DNA, 1:10 dilution (positive control)
• H
2
O (negative PCR control)
Note: When taking aliquots of your samples, always pipette from the center of the coloured phase as an
additional precaution to avoid carrying over remaining break solution.