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. Author manuscript; available in PMC: 2024 Dec 13.
Published in final edited form as: Nat Protoc. 2024 Feb 7;19(5):1381–1399. doi: 10.1038/s41596-024-00953-9

Fig. 3 ∣. Design principles for the implementation of MACHETE.

Fig. 3 ∣

a, A schematic of a generic donor to be integrated into a region of interest. In green is highlighted the selected KI spacer sequence, and the design of the 40-bp homology arms is shown. b, The sequence of the oligonucleotides that are needed to make the targeting sgRNA and the PCR donor to the region highlighted in a. c, Available MACHETE cassettes. d, A schematic of the sequences that will define the intended deletion breakpoint (top) and the control cassette excision (bottom). In purple are highlighted the flanking spacer sequences, and in gray and dark red are highlighted the synthetic spacers to excise the MACHETE cassette. e, The sequence of the oligonucleotides that are needed to make the target and control deletion sgRNAs shown in d. f, Overview and schematic of the MACHETE protocol. QC, quality control. Image c reproduced from ref. 14, Springer Nature Ltd.