Fig. 1

The IMPLANT protocol. In step 1, transgenic plants are generated that contain the gene of interest (GOI) and a selection marker gene with a built-in competitor sequence. This internal sequence can be any sequence, as long it has a similar GC content and can be distinguished from the endogenous gene after amplification with the same primers (indicated in gray) by amplicon size or by size differences after restriction enzyme digestion. For illustrational purposes, we depicted Agrobacterium-mediated transformation with a T-DNA binary vector in a callus-based system with plant regeneration after selection with the marker. In step 2, two different transgenic plants are shown. One plant harbors a single transgene copy (A), while the other contains two transgenes (B). The primer pair is indicated on both the endogenous sequence and the competitor present on the transgene. DNA is harvested from both plants to use in the PCR reaction in step 3. Because the endogenous gene and the transgene will compete for the same primers, the ratio between the peak heights (after capillary electrophoresis) will be proportional to the copy number