Pathways significantly represented in all oocytes and blastocysts.

<p>Listed entries represent transcripts and their respective NCBI accession numbers that were significantly expressed (P<0.05; q≤0.22) in the appropriate sample(s). “-“ denotes no significantly expressed components were detected in the pathway.</p

Pathways significantly represented in oocyte and blastocyst samples with similar molecular signatures.

<p>Listed entries represent transcripts and their respective NCBI accession numbers common to oocyte 1 and 2 relative to oocyte 3 and blastocyst 1 and 2 relative to blastocyst 3. “−” denotes no significantly expressed components were detected in the pathway.</p

Heatmap cluster analysis of individual oocyte and blastocyst gene expression profiles.

<p>Hierarchical clustering was used to compare the gene expression profiles of individual oocytes and blastocysts, with highly expressed genes shown in red, weakly expressed in green. Oocytes show clearly distinct profiles from blastocysts, with oocytes 1 and 2 more similar to each other than to oocyte 3, and blastocyst 1 and 2 more similar to each other than to blastocyst 3. 4-cell embryos were omitted from this analysis based on the low abundance of expressed transcripts.</p

Scattergraph plots comparing gene expression of individual human oocytes, 4-cell embryos and blastocysts.

<p>Each of three individual samples at each stage is compared to each of the others. Red dots represent transcripts called Present in both samples, yellow dots represent transcripts absent in both samples and blue dots represent transcripts Present in one sample and not the other. The innermost oblique lines represent 2-fold differentially expressed transcripts. Additional pairs of lines represent transcripts expressed at 5, 10- and 20-fold, respectively. As expected, fewer genes were called Present at the 4-cell stage which reflects the degradation of polyA containing maternal transcripts by the 4-cell stage during EGA.</p

Primary sequences of aptamers used in the study.

<p>Primary sequences of aptamers used in the study.</p

Scoring system.

<p>Scoring system.</p

Predicted secondary structures and binding affinity to U87MG cells of aptamers SA43 and SA44 DNA.

<p><b>A</b> represents similar secondary structures of SA43 and SA44 aptamers predicted by the M-fold software (Zuker, 2003). Dashes between nucleotides indicate Watson-Crick base pairing. <b>B</b> represents binding affinity of aptamers SA43 and SA44 to live U87MG cells analysed by flow cytometry. The mean fluorescence intensity (MFI) of the cells was plotted against varying concentrations of the Cy3 labelled aptamers (0.5–200 nM) and analysed using non-linear regression analysis.</p

Subcellular localisation of SA43 and SA44 with endoplasmic reticulum (ER), golgi apparatus (GA) and lysosomes.

<p>Live U87MG cells were incubated separately with Cy3 labelled SA43, SA44 and RA at a concentration of 100 nM for 24 hours. CellLight ER and GA labelled with GFP, and lysotracker green DND-26 were then incubated to the cells to track the co-localisation of aptamers with ER, GA and lysosomes respectively, and fixed using 4% PFA. The nuclei were counterstained with DAPI (blue). Cells with random aptamer and marker alone were used as control. Bar = 10 μm.</p

Effect of SA43 and SA44 DNA on cell viability using PrestoBlue assay.

<p>Live cells U87MG, 1321N1 and SVGp12 were incubated separately with SA43 and SA44 aptamer at concentrations of 20 nM, 100 nM, 500 nM and 1000 nM at 37°C (5% CO₂) and assessed for proliferation activity at 24, 48 and 72 hours using the PrestoBlue assay. Cisplatin at 10 μM concentration was used as positive control. A two-way ANOVA test was performed to analyse the difference in the percentage cell viability of the SA43 and SA44 against cisplatin and untreated controls. <b>A</b>, <b>B</b>, and <b>C</b> represent no statistical difference between the percentage cell viability of SA43 and untreated controls for U87MG, 1321N1 and SVGp12 cells, respectively (<i>p</i> > 0.05). <b>D</b>, <b>E</b>, and <b>F</b> represent no statistical difference between the percentage cell viability of SA44 DNA and untreated controls for U87MG, 1321N1 and SVGp12 cells, respectively (<i>p</i> > 0.05). Cisplatin showed a significant decrease in the percentage cell viability at 72 hours when compared to SA43 and SA44 and untreated control (* <i>p</i> < 0.05). All experiments were repeated at least three times. Data are mean of three independent samples. Error bars = Standard deviation.</p

Aptohistochemistry analysis of non-cancerous and different pathological grade glioma patients stained with biotinylated DNA aptamers.

<p><b>Panel X</b> Representative images of tissue sections from non-cancerous and different pathological grade glioma patients stained with biotinylated SA43, SA44 and random aptamer (RA). <b>A-E</b> SA43 treated; <b>F-J</b> SA44 treated; <b>K-N</b> RA treated. <b>Panel Y (a)</b> Scatter plot showing the distribution of average scores for each aptamer on all patient tissue sections. Using ANOVA and post hoc Bonferroni test, SA43 showed statistical difference in the average binding scores between all pathological grades (I, II, III, IV) of glioma tissues compared to the non-cancerous (NC) tissues (*, <i>p</i> < 0.05; **, <i>p</i> < 0.01). <b>Panel Y (b)</b> Fishers exact test confirmed significant difference in binding selectivity between non-cancerous group and tumour groups with grade I, II, III, and IV treated with SA43 aptamer (<i>p</i> < 0.05) (highlighted in bold). SA44 showed no statistical difference in the average binding scores between tumour and NC tissues. <b>Panel Z</b> SA43 showing cell type selectivity within the tissue. Endothelial cells are indicated by arrows in grade I glioma (<b>a</b>), grade II glioma (<b>b</b>), grade III glioma (<b>c</b>), and grade IV GB (<b>d</b>) and show a mix of positive (dash arrows) and negative (straight arrows) staining with SA43 aptamer. <b>e</b> represents negligible binding to Purkinje cells (straight black arrow). All scale bars, 200 μm.</p