sgRNA array

Pairs of ssDNA oligonucleotides ≤200 nt long that encode the necessary genetic parts (promoter, sgRNA, terminator) were ordered from Integrated DNA Technologies (IDT). These oligos are annealed by PCR using KAPA HiFi MasterMix (KAPA Biosystems, #07958935001) and the resulting dsDNA modules were then assembled in a one-pot Golden Gate assembly reaction using type II enzymes BsaI (New England Biolabs, #R0535S) or BsmbI (New England Biolabs, #R0580S) to generate plasmids with different numbers of sgRNAs. After transformation, these plasmids were re-purified and digested with restriction enzyme BsphI (New England Biolabs, #R0517S) to make sure they have the expected sizes and thus rule out the possibility of unwanted homologous recombination during construction and transformation (Supplementary Figure S8).