Part:BBa_K1442034

HCV 3

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. Please refer to Section: RdRP function for further details. The RdRP chosen for our project is taken from the Hepatitis C virus (HCV). It starts replication at the 3’ end of the HCV RNA strand. This 3’ UTR is a RNA promoter. It possesses 3 main features that are essential for supporting RNA replication . First, the poly (U/UC) tail which is present in all HCV strains and is believed to bind the HCV RdRP – NS5B. Second, the presence of the full length 3’X region which has complex stem loop secondary structures that bind cellar and viral factors and facilitate the replication. And third, the 3’ UTR variable region is important as it improves efficiency several times. Thus, the whole length of the promoter is about 230 nt long.

Usage

Used as an RNA promoter to direct replication by RdRP.

RdRP-directed Replication

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

2. The active RdRP binds to the 3’ end of the transcribed plus –sense RNA strand. It produces its complementary minus-sense RNA strand.

3. The RdRP interacts with the 5’ end of the replicated minus-sense RNA strand. It produces the plus-sense RNA strand again and completes the full replication of the RNA. The new RNA strand can act as a template as well and the process of replication can start over.

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
  INCOMPATIBLE WITH RFC[10]

  Illegal PstI site found at 200
  Illegal PstI site found at 227
  
• 12
  INCOMPATIBLE WITH RFC[12]

  Illegal NheI site found at 169
  Illegal PstI site found at 200
  Illegal PstI site found at 227
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal PstI site found at 200
  Illegal PstI site found at 227
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal PstI site found at 200
  Illegal PstI site found at 227
  Illegal NgoMIV site found at 241
  Illegal NgoMIV site found at 270
  
• 1000
  COMPATIBLE WITH RFC[1000]