Fig. 1.

Schematic diagram depicting the workflow comparison between traditional and LION approaches for constructing gRNA expression vectors. a The traditional gRNA vector construction comprises Golden Gate Assembly, transformation of E. coli competent cells, bacterial colonies PCR screening, positive colony expansion by incubating at 37 °C and bacterial culture overnight, and plasmid prep before the CRISPR vector is ready for the cell transfection and activity validation. The whole traditional procedure takes approximately 2.5 days. b The LION method requires only two steps: Golden Gate Assembly and quick DNA purification to generate transfection-grade CRISPR plasmids. The whole procedure takes approximately 5 h (~ 4 h for GGA and less than 1 h for DNA purification