Fig. 5

Real-time PCR analysis of NOPE oligos efficiency. Results of the 3rd PCR are shown. Typical results are shown for the two independent experiments each performed in two replicas. See Table 1 for all oligonucleotides used. a Four different NOPE oligos (NOPE-R0, R1, R2, R3) performance in amplification of EGFR gene exon 20 fragment. Dotted lines represent positive control reaction containing NOPE oligo and two amplification primers: forward TruSeq-short and reverse EGFR-ex20-R1 in the 1st PCR. Solid lines represent test reactions containing NOPE oligo and only reverse primer EGFR-ex20_R1 in the 1st PCR. b Comparison of EGFR-NNN-ex20 primer exclusion efficiency by the four NOPE oligos. dCt between control reaction with two amplification primers in the 1st PCR and test reaction with only reverse primer in the 1st PCR reflects efficiency of EGFR-NNN-ex20 primer exclusion. c Comparison of the NOPE oligos inhibitory effect on amplification of target EGFR gene exon 20 fragment. dCt between amplification with two EGFR exon 20 amplification primers with and without NOPE oligo reflects inhibition of the target DNA amplification by NOPE oligo. d NOPE oligos performance in amplification of EGFR gene exons 18, 19, 21, and 22 fragments. Dotted lines represent control reactions containing NOPE oligo and two amplification primers in the 1st PCR: TruSeq-short and complementary EGFR primer. Solid lines represent test reactions containing NOPE oligo and a single EGFR primer in the 1st PCR. e EGFR-NNN primers exclusion efficiency by NOPE oligos for EGFR gene exons 18, 19, 21, and 22 fragments. dCt between control reaction with two amplification primers in the 1st PCR and test reaction with only the reverse primer in 1st PCR reflects efficiency of of EGFR-NNN primer exclusion. f NOPE oligos inhibitory effect on amplification of target fragments of EGFR gene exons 18, 19, 21, and 22. dCt between reactions containing NOPE oligo and two EGFR amplification primers and reactions containing only EGFR amplification primers reflects inhibition of the target DNA amplification by NOPE oligo