SELEX: barcodes of sequencing primers and numbers of nested PCR cycles used to attach sequencing primers (see Table 4 for sequences of sequencing primers).

<p>SELEX: barcodes of sequencing primers and numbers of nested PCR cycles used to attach sequencing primers (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100572#pone-0100572-t004" target="_blank">Table 4</a> for sequences of sequencing primers).</p

Structure of the combinatorial library that was screened for bivalent aptamers.

<p>Structure of the combinatorial library that was screened for bivalent aptamers.</p

Sequences of aptamers investigated in affinity and inhibition assays; the sequences of APT-15 and APT-29 in bivalent aptamers are underlined.

<p>Sequences of aptamers investigated in affinity and inhibition assays; the sequences of APT-15 and APT-29 in bivalent aptamers are underlined.</p

SELEX: Concentrations of DNA, magnetic beads and number of additional PCR cycles.

<p>SELEX: Concentrations of DNA, magnetic beads and number of additional PCR cycles.</p

Times at which supernatants were collected in single-step selection, barcodes of sequencing primers (see Table 4) that were used to amplify DNA from these supernatants, and number of nested PCR cycles (<b><i>n</i></b>) used to attach primers (see Table 4 for sequences of sequencing primers).

<p>Times at which supernatants were collected in single-step selection, barcodes of sequencing primers (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100572#pone-0100572-t004" target="_blank">Table 4</a>) that were used to amplify DNA from these supernatants, and number of nested PCR cycles (<b><i>n</i></b>) used to attach primers (see <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0100572#pone-0100572-t004" target="_blank">Table 4</a> for sequences of sequencing primers).</p

Bivalent Aptamer Structures.

<p><b>a)</b> Predicted structure of bivalent aptamer based on Linker 2. <b>b)</b> Structures of truncated derivatives of bivalent aptamer based on linker 2; excised bases are shown as grey circles.</p

Bar graphs showing enrichment of motifs 1–3 in each cycle of single-step selection, and motifs 4–6 in each round of SELEX.

<p>Bar graphs showing enrichment of motifs 1–3 in each cycle of single-step selection, and motifs 4–6 in each round of SELEX.</p

Extraction of aptamer sequences from NGS reads.

<p><b>a)</b> Informatics pipeline as applied to DNA retained on the beads in single-step selection. Key: numbers in rectangles show the number of sequences at each stage in the pipeline. <b>b)</b> Linkers 4–6 identified in round 5 of SELEX.</p

Workflow of single step selection.

<p><b>a)</b> Key: size selection  =  preparative electrophoresis; QC  =  quality control; NGS  =  next generation sequencing. <b>b)</b> Detail of one cycle of dissociation. Thrombin-coated magnetic beads (MBs) with bound DNA are incubated for time <b>t</b> and then magnetically precipitated. DNA in the supernatant was extracted and fed into PCR I, and the MBs were re-suspended for a new cycle of dissociation. <b>c</b>) Bar-chart showing durations (<b>t</b>) of the dissociation cycles.</p

IC₅₀ values, affinities and base pairing between scaffold and linker in bivalent aptamers.

<p>IC₅₀ values, affinities and base pairing between scaffold and linker in bivalent aptamers.</p