Fig. 2

Identification of circAnks1a. a Scheme illustrating the production of circAnks1a. CircAnks1a was formed by back splicing from exon 5 to exon 11 of the Anks1a gene. b The expression of circAnks1a was validated by Sanger sequencing. The arrow indicates the junction site of circAnks1a. c Random hexamer or oligo(dT)₁₈ primers were used in the reverse transcription experiments. The RNA levels were analyzed by qPCR and normalized to the value obtained using random hexamer primers (^**P < 0.01 vs. random hexamer primers group, two-tailed two-sample t tests, n = 4). d PCR showed that dorsal horn circAnks1a, but not mAnks1a, was resistant to digestion by RNase-R (n = 3). e Northern blotting showed that circAnks1a was detected by circAnks1a probes after RNase R treatment, while mAnks1a was not (n = 3). f The relative RNA levels of circAnks1a and mAnks1a in the spinal dorsal horn were analyzed by qPCR at various time points after treatment with actinomycin D (i.t.) (^**P < 0.01 vs. the mAnks1a group at 0 h, ^##P < 0.05 vs. the circAnks1a group at 0 h, two-tailed one-way ANOVA, n = 5). g The expression of circAnks1a in nine different tissues was examined in naïve, sham and SNL-treated rats (^**P < 0.01 vs. the sham group, two-tailed one-way ANOVA, n = 6). h FISH showed that circAnks1a was expressed in the nucleus (arrow) and cytoplasm (arrowhead) of spinal dorsal horn neurons. Left scale bar, 50 μm; right scale bar, 10 μm (n = 3). i PCR results obtained after nucleus and cytoplasm separation showed circAnks1a was expressed in both the nucleus and the cytoplasm (n = 5). The data are presented as the mean ± s.e.m. Source data is available as a Source Data file