Genome Engineering by Transgene- Instructed Gene Conversion in C. elegans - Caenorhabditis Elegans: Molecular Genetics and Development - page 76

Fig. 4 Jumping PCR. (A) Example of jumping PCR occurring during the screening for a KI allele. A PCR product can be
generated by ''jumping'' between two single-strand DNA products synthesized from the transgenic repair template and the
genome respectively. This will generate a false-positive PCR signal even in the absence of a MosTIC-engineered allele (adapted
from Robert et al., 2009 ). For primers names, we use the same nomenclature as in Fig. 3 . (B) Effect of annealing temperature on
jumping PCR. In this experiment, mixed transgenic/nontransgenic worm populations derived from four non-heat-shocked
independent lines (a, b, c, d) constructed to engineer by MosTIC a KI allele at the unc-5 locus were analyzed by PCR using
the primers designed for the MosTIC allele PCR screening. At annealing temperatures (indicated at the top of the gels) ranging
from 60 to 66.4 C, false-positive PCR products are generated by jumping PCR. They disappear at 68.4 and 70 C. Two dilutions
of a control plasmid (P1=1 ng/ m L and P2=10 -2 ng/ m L) are used to verify that the amplification is still efficient in the presence of
a specific PCR template. (C) Example of a PCR screen performed on animals derived from the lines tested in (B). Mixed
transgenic/nontransgenic populations derived from heat-shocked transgenic animals are tested by PCR. When primer annealing
is performed at 64 and 66.4 C, many false-positive signals are detected. When annealing is performed at 68.4 C, only one signal
is present (pool #3). Further sibling selection experiments performed on pool 3 identified worms carrying the desired MosTIC-
engineered unc-5 locus.