Fig. 4

LbCas12a/crRNA RNP complex is an efficient and robust tool for short DNA fragment deletion and targeted gene disruption in X. tropicalis. a PCR assay data show segmental deletions induced by LbCas12a/crRNA RNP complex with paired crRNAs at the tyr locus of X. tropicalis embryos. M, DNA molecular weight marker. crRNAs T1 pairs with T2 and T3 pairs with T4. b Sanger DNA sequencing data show segmental deletions as well as indels induced by LbCas12a/crRNA RNP complex with paired crRNAs at the tyr locus. For both panels, the wild-type sequence is shown at the top with the target sites in red and the PAM sequence in blue. In the upper panel, 120 bp wild-type sequences were not shown. Red dashes indicate deletions and lowercase letters in red indicate insertions or mutations. The numbers in parentheses represent the ratio of this sequence in the total colonies sequenced. c T7EI assay data show targeted mutations induced by ptf1a/p48, sftpb, tbx5, or ctcf crRNAs and the LbCas12a RNP complexes. M, DNA molecular weight marker. d Sanger DNA sequencing data show targeted mutations induced by ptf1a/p48, or tbx5 crRNAs and the LbCas12a RNP complexes. For both panels, the wild-type sequence is shown at the top with the target sites in red and the PAM sequence in blue. Red dashes indicate deletions and lowercase letters in red indicate mutations. The numbers in parentheses indicate the ratio of this sequence in the total colonies sequenced