Part:BBa_K2323004

Lwa Cas13a under T7 promoter with solubility tag

Uses a T7 promoter and RBS B0034 (BBa_K525998) to express Lwa Cas13a (BBa_K2323000) flanked by a 6x His-Tag to allow purification and a SUMO-tag to improve solubility. This is terminated by a Tphi terminator. The whole coding sequence is also available as BBa_K2323002.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 1768
  Illegal BglII site found at 3448
  Illegal BamHI site found at 144
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 2847
  

Usage and Biology

Cas13a from Leptotrichia wadei (Lwa) is class 2 type VI CRISPR-Cas effector. It is capable of targeting specific ssRNA and cleaving them using its bounded crRNA as guide strand. Unlike other ssRNA CRISPR-Cas effectors, Cas13a is capable of processing the precursor crRNA (pre-crRNA) by itself, cleaving, binding the RNA sequence and then building the Cas13a:crRNA surveillance complex. One characteristic of Cas13a along with other type VI members is its ability to degrade other RNA starnds regardless of their sequences, provided the protein has been activated by binding and digesting the target sequence complementary to the crRNA. This "collateral activity" makes Cas13a a powerful biological tool, as unspecific RNA degradation can be triggered on command by providing the target RNA.