Fig. 4From: Optimized PCR conditions minimizing the formation of chimeric DNA molecules from MPRA plasmid librariesComparison of PCR approaches in terms of proportion of chimeric products generated per BC from MPRA libraries. BC–ROI regions of library-71 and library-83 were amplified by emulsion and conventional two-round PCR using the same settings (10 pg of template DNA, 15 amplification cycles and elongation time of 30 s in round #1 PCR, 1/100th of the purified products of round #1 PCR as template, 18 amplification cycles and elongation time of 30 s in round #2 PCR) and subsequently subjected to NGS. The experiments were done in duplicates and averaged values of proportion of chimeric products per BC are plotted as histograms separately for library-71 (a) and library-83 (b). Pronounced peaks at middle values of proportion of chimeric products per BC are mainly a result of low NGS coverage of some BCs (i.e., the peaks primarily represent cases with 1 chimeric BC–ROI combination per several genuine ones)Back to article page