Coding

Part:BBa_K559000:Design

Designed by: Jacky, Fong Chuen, Loo   Group: iGEM11_Hong_Kong-CUHK   (2011-09-29)
Revision as of 21:14, 14 October 2011 by Jacky Loo (Talk | contribs) (→‎Source)

Halorhodopsin


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal XhoI site found at 280
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 292
    Illegal AgeI site found at 18
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI site found at 228



Design Notes

N.pharaonis was kindly given as a gift from I. P. Hohenfeld, who is an expert studying halorhodopsin [1]. The halorhodopsin gene was constructed by PCR and linked with RFC[10] prefix and suffix.

Template

pET-27b-HRNegre03

  • The DNA sequence selected from ID J05199.1 of 61nt to 700nt for functional protein coding [2].
The figure illustrates our work to prepare HR gene from bacterial gene to biobrick backbone.

Forward primer

5'- CGG AAT TCG CGG CCG CTT CTA GAG ATG GCT GAG ACA TTG CCA CCG G -3'

Reverse primer

5'- CGC TGC AGC GGC CGC TAC TAG TAC TTA TCA GTG GTG GTG GTG GTG GTG GCG TAT TGA ACA TAT CCG C -3'

Source

  • HR gene from N.pharaonis (GenBank ID: J05199.1)

Acknowedgment

The gene of this biobrick Halorhodopsin came as a gift from P. Hohenfeld, A. A. Wegener,M. Engelhard (1999): Purification of histidine tagged bacteriorhodopsin, pharaonis halorhodopsin and pharaonissensory rhodopsin II functionally expressed in Escherichia. FEBS Lett.

References

[1] Hohenfeld, I.P. “Purification of histidine tagged bacteriorhodopsin, pharaonis halorhodopsin and pharaonis sensory rhodopsin II functionally expressed in Escherichia coli,” FEBS Letters, vol. 442, Jan. 1999, pp. 198-202. [2] Lanyi, J.K. Halorhodopsin, a light-driven electrogenic chloride-transport system. Physiological reviews 70, 319(1990). [3 Essen, L.O. Halorhodopsin: light-driven ion pumping made simple? Current opinion in structural biology 12, 516–522(2002)]