Part:BBa_K3286012

dCas9

Usage and Biology

This part is the catalytically inactive version of Streptococcus pyogenes. It is identical to Part:BBa_K2558003 apart from an extra 41 nucleotides at the 5' UTR. The characterization of this part is also found under Part:BBa_K2558003 or as part of an Acr-dCas9 gene circuit under Part:BBa_K3286010.

CRISPR interference (CRISPRi) makes use of catalytically inactive variants of Cas9 (dCas9) proteins to suppress gene expression [1]. Identical to their active counterparts, the co-expression of guide RNAs directs the ribonuclease protein (RNP) to its specific DNA target sequence. However, introduction of mutations in the RuvC1 and HNH nuclease domains of Cas9 cause the Cas9 to lose endonuclease activity, without impeding the DNA binding [2; 3]. This enables the reversible transcriptional inhibition by tightly DNA-bound dCas proteins, contrary to irreversible cleavage by active Cas9.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 1140
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BamHI site found at 3419
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]