Genome Editing

CRISPR Gene Editing

  1. G+FLAS
  2. Genome Editing
  3. CRISPR Gene Editing

CRISPR gene editing technology is a power tool for editing genomes, which originated from an antiviral defense system of bacteria. CRISPR stands for “clusters of regularly interspaced short palindromic repeats.” CRISPR gene editing technology uses RNA-guided endonucleases to precisely locate and cut target nucleic acid sequences. The RNA-endonuclease complex consists of CRISPR-associated protein, Cas, that binds and cleaves nucleic acids, and a strand of RNA that guides Cas to the exact location in genome.

Class 2 Cas Proteins

Type II
Cas9
Nuclease Domain RuvC, HNH
tracrRNA present
PAM GC-rich sequence at 3’
Substrate double-stranded DNA
Cutting Mechanism blunt end cuts

Type V-A
Cas12a (Cpf1)
Nuclease Domain RuvC, Nuc
tracrRNA absent
PAM AT-rich sequence at 5’
Substrate double-stranded DNA
Cutting Mechanism staggered end cuts with 5’ overhangs

Type V-B
Cas12b (C2c1)
Nuclease Domain RuvC
tracrRNA present
PAM AT-rich sequence at 5’
Substrate double-stranded DNA
Cutting Mechanism staggered seven-nucleotide cut of target DNA

Type VI-A
Cas13a (C2c2)
Nuclease Domain 2 HEPN domains
tracrRNA absent
PAM PFS w/o G at 5’
Substrate single-stranded RNA
Cutting Mechanism cleaves ssRNA near uracil and has collateral activity

Type VI-B
Cas13b (C2c3)
Nuclease Domain 2 HEPN domains
tracrRNA absent
PAM 5’, w/o C, 3’, NAN/NNA of PFS
Substrate single-stranded RNA
Cutting Mechanism cleaves ssRNA near C, U and has collateral activity

Redrawn from Shmakov et al. Nat. Rev. Microbiol. 2017
Smargon et al. Mol. Cell. 2017