Part:BBa_K1442108

C2HP RdRP Promoter regulated by MS2

Each unique RNA dependent RNA polymerase (RdRP) initiates de novo replication of a RNA strand by interacting with RdRP-specific RNA sequences, henceforth called RdRP/RNA promoters. The RdRP chosen for our project is taken from the Hepatitis C virus (HCV). It stops replication at the end of the RNA template. The C2HP RNA terminator was designed by us based on the C2 promoter identified by Heinz, Kao (2000) . A MS2 Box (BioBrick) was put in front of the promoter sequence to test if its secondary structure would be enough to disturb the interaction between the RdRP and the promoter. When incorporated in the full Replicon system, the MS2 protein will also bind to the MS2 box and interfere with the RdRP. Please refer to Section: RdRP-directed Replication and C2HP for further details.

Usage

Used as an RNA promoter that could be dowregulated by the interaction of a MS2 domain that acts as a "transcription factor", where the MS2 interferes with direct replication by RdRP.

RdRP-directed Replication and C2HP

1. The DNA strand gets transcribed and then translated, whereby the RdRP protein is produced.

2. The active RdRP cannot bind to the 3’ end of the transcribed plus –sense RNA strand due to the Hairpin (HP). It does not produce its complementary minus-sense RNA strand. Replication does not occur.

Mathematical Modelling

E-coli

Where c, α-,β are constants, E stands for the RdRP, G – for GFP. R- is amount of minus-sense RNA strands, R+ amount of plus-sense RNA strands; µ- is degradation rate of the minus-sense strands and µ+of the plus-sense.

Sequence and Features

Assembly Compatibility:

• 10
  COMPATIBLE WITH RFC[10]
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal NgoMIV site found at 57
  Illegal NgoMIV site found at 86
  
• 1000
  COMPATIBLE WITH RFC[1000]