Part:BBa_K2683017

Release Factor 2 in pSB1C3

Release Factor 2 (RF2) is a protein that causes termination of translation by recognizing the stop codon in an mRNA sequence. This part was originally designed by Lethbridge iGEM team 2017(http://2017.igem.org/Team:Lethbridge) The original composite part can be found here: https://parts.igem.org/Part:BBa_K2443030 RF2 has been Codon optimized for use in Escherichia coli and contains a N-Terminal hexahistidine tag. It is under the regulation of T7 Promoter, RBS and double terminator.

We have improved the part by successfully inserting it into the pSB1C3 standard plasmid.

Figure 1- The overexpression of Release Factor 2 in pSB1C3 as run on a 12% SDS-PAGE gel. Samples taken correspond to point of induction and samples taken over the next three hours. The protein is expected at about 43kDa.

Figure 2- 12% SDS-PAGE showing the nickel affinity chromatography purification of Release Factor 2. Elutions 1-5 show the purified proteins at the 50kDa range.

Sequence and Features

Assembly Compatibility:

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

  Illegal BglII site found at 659
  Illegal BglII site found at 1116
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]