Part:BBa_K2047010:Experience

_NOTOC__ Fluorescent reporter system with stem-loop, GFP_Stem-loop with free energy of -25.6 kcal/mol

Assembly Compatibility:

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

  Illegal NheI site found at 764
  
• 21
  COMPATIBLE WITH RFC[21]
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  INCOMPATIBLE WITH RFC[1000]

  Illegal BsaI.rc site found at 644
  

Design Notes

none When stem loops inserted in the 3’ end of the upstream gene, it protects its mRNA against the cleavage of exonuclease, increasing the ratio of abundance of the first gene product relative to that of the second gene product. Furthermore, the lower free energy of stem loops are, the more stable the secondary transcripts of the upstream are, tuning the expression of multiple genes. We designed a series of stem loops with different free energy and measured it by fluorescence reporter system.

Source

Inspired by Xu’s work and based on keasling’s work, we designed a series of stem loops with different free energy for further use as basic regulatory parts.

References

[1]. Pfleger, B.F., et al., Combinatorial engineering of intergenic regions in operons tunes expression of multiple genes. Nat Biotechnol, 2006. 24(8): p. 1027-32.
[2]. Xu, C., et al., Cellulosome stoichiometry in Clostridium cellulolyticum is regulated by selective RNA processing and stabilization. Nat Commun, 2015. 6: p. 6900.
[3]. Smolke, C.D. and J.D. Keasling, Effect of gene location, mRNA secondary structures, and RNase sites on expression of two genes in an engineered operon. Biotechnol Bioeng, 2002. 80(7): p. 762-76.