Part:BBa_K598026

AND gate (BBa_K228258) regulated by TPP riboswitch 1.20 (BBa_K598004)

This is an AND gate regulated by a TPP riboswitch. The translational strength of T7ptag is regulated by thiamine pyrophosphate concentration.

The AND gate we utilized was designed by Anderson and his colleagues and consequently standardly redesigned by [http://2009.igem.org/Team:PKU_Beijing/Project/AND_Gate_1/Design PKU_Beijing 09 team].

Figure 1: The construction and mechanism of AND gate. A). The standardly redesigned AND gate of PKU_Beijing 09 team. B). The topology and mechanism of AND gate. Parts showing in box constitute the core processing module. .

As shown in Fig. 1B, pBAD promoter serves as input 1. It is activated by arabinose, leading to the expression of SupD, an amber suppressor tRNA. The input 2 is pSal promoter activated by salicylate via binding to NahR. It regulates the expression of a T7 polymerase coding sequence with two amber codons reside in the position 8 and 14 (T7ptag), resulting in premature translational termination. When both T7ptag and SupD tRNA genes are expressed, functional T7 polymerase would be synthesized and consequently actives output, T7 promoter. A green fluorescent protein (GFP) generator regulated by T7 promoter serves as a reporter.

By placing a TPP-responsive RNA controller element upstream of the coding sequence, we obtained an AND gate modulator whose T7ptag gene translation rate varies in response to ligand concentration.

Ligand responsive pattern of AND gate. .

Sequence and Features

Assembly Compatibility:

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

  Illegal NheI site found at 1274
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 2144
  Illegal BamHI site found at 1214
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 1976
  Illegal AgeI site found at 1320
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI site found at 1323