Part:BBa_K4472988:Design

Split ribozyme detecting hcat and expressing eforRED

Assembly Compatibility:

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

  Illegal NheI site found at 7
  Illegal NheI site found at 30
  Illegal NheI site found at 331
  Illegal NheI site found at 354
  Illegal NheI site found at 686
  
• 21
  INCOMPATIBLE WITH RFC[21]

  Illegal BglII site found at 588
  Illegal XhoI site found at 135
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Design Notes

The promoters were to weak to get a signal in our assays, so we suggest to anyone else to use much strogner promoters. The promoters of split ribozyme half 1 and split ribozyme half 2 need to be different in order to get them synthesized. If they are the same there is too much repetition for the synthesis companies. The guideRNA should have a length of 82 basepairs (Gambill L, Staubus A, Ameruoso A, Chappell J. A split ribozyme that links detection of a native RNA to orthogonal protein outputs. bioRxiv. January 2022:2022.01.12.476080. doi:10.1101/2022.01.12.476080)

Source

The original ribozmye is from Tetrahymena thermophila. The guide RNA is complementary to the stable endogenous E.coli mRNA hcat.

References

Gambill L, Staubus A, Ameruoso A, Chappell J. A split ribozyme that links detection of a native RNA to orthogonal protein outputs. bioRxiv. January 2022:2022.01.12.476080. doi:10.1101/2022.01.12.476080

Zhou K, Zhou L, Lim Q, Zou R, Stephanopoulos G, Too HP. Novel reference genes for quantifying transcriptional responses of Escherichia coli to protein overexpression by quantitative PCR. BMC Mol Biol. 2011;12(1):1-9. doi:10.1186/1471-2199-12-18/FIGURES/5

https://www.ncbi.nlm.nih.gov/nuccore/NC_000913.3?from=2666707&to=2667846&report=genbank&strand=true