Figure 3
From: Cell-Free DNA Kinetics in a Pre-Clinical Model of Head and Neck Cancer
Detection and quantification of CRPV DNA in VX2 cells. (a) Flow diagram of the iterative process of optimizing and selecting a sufficiently specific and sensitive qPCR assay for detection of CRPV DNA. (b) Map of the CRPV genome highlighting the E6 open reading frame and the sets of qPCR primers tested during optimization process referred to in (a). Beige boxes indicate sets of primers with the same 5′ start site but differing in amplicon size. (c) Plot of qPCR cycle threshold (Ct) vs. amount of VX2 DNA for CRPV E6 qPCR assay. (d) Melt curve for CRPV E6 qPCR assay (red) showing no signal in uninjected rabbit plasma (blue) (n = 4 rabbits). (e) Quantification of copy number of CRPV genome in VX2 rabbit tumour line by multiplexed ddPCR. 2-D plot of droplet fluorescence for FAM and HEX fluorophores from probes for CRPV E6 gene and rabbit ACTA2 probes respectively. The CRPV copy number per diploid genome: 29.3 ± 1.3. Error bars represent SD.
