Shuffling the yeast genome using CRISPR/Cas9-generated DSBs that target the transposable Ty1 elements
Fig 3
Sequence of CRISPR/Cas9 target in the Ty element, and diagnosis of events causing loss of URA3 insertion from Ty1::URA3.
(A) Sequence in Ty1 targeted by the guide RNA (shown in red). (B) Position of marked Ty element on chromosome III, and location of PCR primers used to diagnose 5-FOA-resistant isolates. The distance between the DSB site (CRISPR/Cas9 target) and the 3’ end of URA3 is about 2.5 kb. Large black arrows show the LTRs (delta elements) associated with Ty1. (C) Different classes of 5-FOA-resistant isolates based on PCR analysis. The sizes of the fragments resulting from PCR reactions with various combinations of primers are given in S3 Table. The wavy line in Class 4 indicates a different flanking sequence from the original Ty1 element. (D) CHEF gel analysis of rearranged chromosome III in MD745-21F. The size of chromosome III was altered as a result of a crossover-associated gene conversion. MD745 is the control strain.