Fig. 3

Characterization of the circRNAs expressed in the mouse preimplantation embryos. a The number of circRNA transcripts and the number of reads mapping to circRNAs (only counting the reads spanning the end-joining site of circRNAs) per million mapped paired-end reads at each stage of mouse preimplantation embryos. Five biological replicates were used for the oocyte samples, and three biological replicates for each of the other six stages. The concentration of circRNAs is elevated between the two-cell and eight-cell stages and decreased drastically at the morula and blastocyst stages. b The expression dynamics of five circRNAs analyzed by RT-qPCR. Top: circRNAs generated from the same gene share the 5′ exon to form the head-to-tail junction sites. The arcs show the joint between two exons. The numbers indicate the junction reads for corresponding circRNAs in the SUPeR-seq dataset. CircRNAs with longer downstream introns show higher expression levels as measured by the circRNA-specific end-joining reads. Middle: RT-qPCR of the linear transcripts from the mouse oocyte to blastocyst stage. Bottom: the abundance of circRNAs analyzed by RT-qPCR from the mouse oocyte to blastocyst stage. GFP RNAs were spiked-in during purification of total RNAs as a control for the technical variation in each sample. Ezh2-circRNA stands for chr6:47540677–47577667 (exons 2–15 of Ezh2), Cntln-circRNA stands for chr4:84971131–85006524 (exons 6–12 of Cntln), Akap7-circRNA#1 stands for chr10:25283892–25289730 (exons 2–3 of Akap7), Akap7-circRNA#2 stands for chr10:25267307–25289730 (exons 2–5 of Akap7), Akap7-circRNA#3 stands for chr10:25220610–25289730 (exons 2–7 of Akap7). The circRNAs are more stable compared with linear ones during the maternal to zygotic transition. c The relationship of circRNA read counts with the host gene FPKM. Genes detected with higher circRNA counts also show higher expression levels of linear transcripts. d Length distributions of flanking introns of the 2891 circRNAs found in mouse preimplantation embryos. The upstream intron is originally adjacent to the first exon of a circRNA before cyclization and the downstream intron is adjacent to the last exon. Both these specific upstream and downstream introns tend to be much longer than other introns in the same gene. e An example (Ccdc171) showing that one host linear RNA gene could generate multiple circRNA species with different joints. All the exons are indicated and numbered along the gene. The arcs between different exons within the circle show end-joining events of the circRNAs