Digital.CSIC. Repositorio Institucional del CSIC

oai:digital.csic.es:10261/311088

A) Genomic map showing positions of genes and locations of mutations from CRISPR/Cas9 mutagenesis in the 9x-saur mutant based on the TAIR10 Arabidopsis genome annotation. The first sgRNA directed cuts in both SAUR61 and SAUR64 (green arrows), creating a deletion between them (green bar) and leaving behind a hybrid gene with a frameshift at the junction site (symbolized by a green X). The second sgRNA directed cuts in the remaining genes (blue arrows, with lighter blue indicating slight mismatches between the sgRNA and the genome), leading to deletions (blue bars) and/or frameshift mutations (blue X’s). SAUR gene names are abbreviated as S61 etc. SAUR61-SAUR68 are on chromosome 1 and SAUR75 is on chromosome 5. B,C) 5-day-old seedlings grown on 1x MS/1% Suc medium in long days. Scale bar, 1 mm. D) Hypocotyl lengths of seedlings grown for 4d in short days on 0.5x MS medium. n, 27 (wild type), 22 (9x-saur). E) Cotyledon area of seedlings grown on vertically oriented plates for 6d on MS/1% Suc medium. n, 16 (wild type), 22 (9x-saur). Graphs show means ± s.d. No statistical differences were detected between wild-type and 9x-saur mutant measurements by t-test. F) Sequences of guide RNAs used for CRISPR/Cas9-mediated mutagenesis, wild-type genes, and mutant alleles present in the 9x-saur mutant. Underlines indicate PAM motif adjacent to guide RNA target site, and any mismatches to the guide RNA sequence. Uppercase bold letters indicate insertion mutations. All alleles create frameshift mutations except for saur75-1, which has an in-frame deletion of 13 amino acids in the SAUR domain., Peer reviewed