Fig. 4: Selective pulldown of double-stranded cDNA for the enhancement of expression profiles. | Communications Biology

Fig. 4: Selective pulldown of double-stranded cDNA for the enhancement of expression profiles.

From: A smart polymer for sequence-selective binding, pulldown, and release of DNA targets

Fig. 4

a The target, a catcher strand library (CSL), and MeRPy are mixed and heated to 95 °C. Subsequently, the sample is cooled to bind catcher strands to the target and MeRPy. MeRPy is quickly precipitated to deplete the target from solution. b dPAGE after pulldown of a dsDNA target (150 bp) with MeRPy-100, showing high pulldown efficiency and specificity. The original uncropped scan of the gel is shown in Supplementary Fig. 12. c Selective depletion of high-abundance insulin (INS), glucagon (GCG), and transthyretin (TTR) cDNA from a clinical NGS library by MeRPy in the presence and absence of an INS-, GCG-, and TTR-targeting CSL. Blue and red data points represent genes with higher and lower transcripts per million (TPM) values, respectively, as compared to the original sample (untreated control). d Pulldown efficiencies for INS, TTR, and GCG, as quantified from RNA-seq (n = 3 independent experiments). e Relative base count after pulldown (blue trace) and standard deviation (n = 3 independent experiments, gray shade), as a function of base position in the transcripts (see Supplementary Note 2). The plot provides single-base-resolution information about depletion efficiencies. CSL-targeted transcript regions are highlighted in beige. Base positions (x-axis values) are relative to the center of the respective targeted region. Dashed lines mark exon boundaries. f Total number of genes detected with >1 TPM in the original sample vs. MeRPy-treated samples (n = 3 independent experiments).

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