IL3000 minicircle annotation and gRNA alignments

We annotated IL3000 minicircles from de novo assembly with a bespoken pipeline and followed the definition for gRNAs from a previous study. We aligned minicircle sequences to known edited mRNAs allowing G-U wobble base pairing to identify high-quality canonical gRNAs encoded within cassettes. The motifs of forward and reverse repeats and the initiation sequence enabled subsequent characterization of cassettes on each minicircle and the detection of non-canonical gRNAs by nucleotide bias. We defined canonical gRNA genes as gRNAs aligned to the edited mRNA without gaps for at least 25 nt with a maximum of three mismatches. The criteria were somewhat arbitrary and may misclassify some gRNAs with parameters close to the cutoffs. However, the strong association of gene characters we observed suggested that the ambiguities were unlikely to sway our conclusion. We detected 556 gRNAs in total. Excluding gRNAs aligned to the non-variable regions on alternatively edited mRNAs gave 465 gRNAs