Part:BBa_K1442116

SLC8 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. The RdRP chosen for our project is taken from the Hepatitis C virus (HCV). The SLC8 RNA promoter was identified as such by Heinz, Kao (2000) , along with a number of other sequences. All of them possess a few common characteristics: an initiation cytidylate at the 3’ end, where the replication starts; and a stable secondary structure- single stranded tail and a stem of various length. Replication stops at the 5’ end. Please refer to Section: RdRP-directed Replication for further details.

This is a genomic plus-strand RNAs that can direct BMV minus-strand initiation and is derived from the 3’ terminal of the Brome Mosaic Virus which forms tRNA like structures. There is a stem loop structure within this, named SLC, that is seen to direct replication. This is also from the BMV SLC loop but has 8 nucleotides added to it which contain the 3’ initiation site CCA.

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 XbaI site found at 33
  
• 12
  COMPATIBLE WITH RFC[12]
• 21
  COMPATIBLE WITH RFC[21]
• 23
  INCOMPATIBLE WITH RFC[23]

  Illegal XbaI site found at 33
  
• 25
  INCOMPATIBLE WITH RFC[25]

  Illegal XbaI site found at 33
  Illegal NgoMIV site found at 57
  Illegal NgoMIV site found at 86
  
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 49