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• Eric-Paul Bennett
• Narimatsu et al

More than 200 glycosyltransferases are involved in biosynthesis of the human glycome, which is comprised of all glycan structures found on different glycoconjugates in all cells. In short of the necessary analytic strategies that enable researchers to decipher the biological roles of any glycan with respect to structure and/or function, the biosynthetic roles of most of these enzymes have been defined. Thus, GlycoCRISPR provides the first collection of CRISPR/Cas9 gRNAs targeting all human glycosyltransferase encoded genes (glycogenes). The uniqueness of this collection becomes evident from the fact that all gRNA designs deposited in the collection have been experimentally validated by recently published methods (Lonowski et al., 2017) to give rise to >30% target cutting efficiency based on in depth indel profiling by IDAA (Yang et al., 2015). GlycoCRISPR gRNAs are made available as a sustainable plasmid format encoding U6 promoter driven gRNA expression. The promised high indel formation capacity of all glycogene gRNA designs enables easy genetic dissection of glycan biosynthetic pathways either as single and/or multiplex combinatorial targeting of the human glycosyltransferase genome.

Workflow for establishment of the GlycoCRISPR collection 205.3 KB

Additional References

• Lonowski LA, Narimatsu Y, Riaz A, Delay CE, Yang Z, Niola F, Duda K, Ober EA, Clausen H, Wandall HH, Hansen SH, Bennett EP, Frödin M. 2017. Genome editing using FACS enrichment of nuclease-expressing cells and indel detection by amplicon analysis. Nat Protoc. Mar;12(3):581-603. PMID: 28207001
• Yang Z, Steentoft C, Hauge C, Hansen L, Thomsen AL, Niola F, Vester-Christensen MB, Frödin M, Clausen H, Wandall HH, Bennett EP. 2015. Fast and sensitive detection of indels induced by precise gene targeting. Nucleic Acids Res. May 19;43(9):e59. PMID: 25753669