Looking to set up a multiplex ddPCR panel for NSCLC driver mutations
from cfDNA - Bio-Rad QX platform. Want to cover the actionable hotspots
in one or two reactions so we can run liquid biopsies efficiently.

Hotspots:
- KRAS G12C (c.34G>T) and G12D (c.35G>A)
- EGFR L858R (c.2573T>G) and T790M (c.2369C>T)
- BRAF V600E (c.1799T>A)
- EML4-ALK fusion, variant 1 (E13;A20)

Plus a wild-type reference for each gene so we can calculate allele
fractions. Pick the right discriminatory strategy per target (ARMS,
LNA-blocking probe, or hydrolysis-probe SNP discrimination - your call,
just justify it in the report).

The make-or-break QC step is multiplex compatibility - primer-dimer and
probe-dimer screening across the full panel, Tm matching, no homopolymer
artifacts. Flag anything borderline rather than forcing it through.

Need a channel-layout plan for both QX200 (2 channels) and QX600
(6 channels), in-silico specificity checks against the human transcriptome
and gnomAD common SNP positions under the probes, and synthetic gBlock
sequences for each mutant and WT target ready to order. Final report
should include the LoB/LoD validation plan.
