Part:BBa_K4140026

Regulatory device

Part Description

The composite component encodes a particular gRNA that is regulated by the human U6 promoter. It is used to direct the Cas12g protein to the target RNA. The interaction of L7Ae with the Kink turn loop inhibits this system, limiting the expression of Cas12g.

Usage

We used this composite as our regulatory system to control the expression of our reporter gene in therapeutic and diagnostic circuit based on CRISPR system which is formed of two component: The first is Cas12g which is a protein that interact differ from other Cas proteins in taking RNA as a substrate by making post transcriptional modification to it that hinders its translation The second part is a gRNA which have affinity to the target RNA sequence needed to be regulated by CRISPR system activity In our system the gRNA is designed to target RNA of PAH to repress it’s translation as shown in figure 1.

Figure 1. (shows the regulatory circuit in our whole cell-based biosensor.)

Sequence and Features

Assembly Compatibility:

• 10
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 820
  Illegal PstI site found at 1294
  Illegal PstI site found at 2236
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal PstI site found at 820
  Illegal PstI site found at 1294
  Illegal PstI site found at 2236
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal XhoI site found at 1815
  
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 820
  Illegal PstI site found at 1294
  Illegal PstI site found at 2236
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 820
  Illegal PstI site found at 1294
  Illegal PstI site found at 2236
  Illegal NgoMIV site found at 1737
  
• 1000
  COMPATIBLE WITH RFC[1000]