Fig. 1

Characteristic of circHAS2 and identification as a biomarker of CRC. A Schematic diagram and heatmap depicting whole-transcriptome RNA sequencing results from three independent replicates of SW620 cells cultured with either vehicle control or anlotinib (10 µM) for 24 h. B Relative expression of top 10 most upregulated and downregulated circRNAs in 5 CRC tumor tissues and paired normal tissues. C Schematic diagram illustrating the formation of circHAS2 from host gene and the back-splice junction site verified by Sanger sequencing. D PCR and agarose gel electrophoresis analysis in SW620 cells verified that the divergent primers for circHAS2 could be amplified from cDNA but not gDNA. E Relative expression of circHAS2 and HAS2 mRNA after treatment with RNase R in SW620 cells. F RNA abundance of circHAS2 and HAS2 mRNA after treatment with Actinomycin D in PCR and agarose gel electrophoresis analysis in SW620 cells. G Representative images of the FISH assay in SW620 cells revealed circHAS2 predominantly located in the cytoplasm with the target probe labeled with Cy3 and nuclei stained with DAPI (scale bar: 20 μm). H Nuclear and cytoplasm fractionation assays in SW620 cells showed the subcellular location of circHAS2. I The expression levels of circHAS2 in CRC tissues and paired normal tissues of microarrays were verified by FISH (scale bar: 100 μm). J The expression levels of circHAS2 in CRC tissues and paired normal tissues of microarrays were confirmed by IHC scores. K The expression of circHAS2 in different TNM stages was detected by FISH (scale bar: 400 μm). L Overall survival curves of 70 CRC patients based on circHAS2 expression levels in our research center. Statistical significance in two-group experiments was evaluated using a two-sided Student’s t-test, and survival analysis was conducted using the Log-rank method. Data are represented as mean ± SD; *, P < 0.05; **, P < 0.01; ***, P < 0.001; ns, no significance