Part:BBa_K1763430:Design

MaSp2-12(1C3)

Assembly Compatibility:

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

  Illegal BamHI site found at 1264
  
• 23
  COMPATIBLE WITH RFC[23]
• 25
  COMPATIBLE WITH RFC[25]
• 1000
  COMPATIBLE WITH RFC[1000]

Design Notes

This construct was assembled using ICA using the spider silk [http://2015.igem.org/Team:UCLA/Notebook/Spider_Silk_Genetics#List_of_Parts ICA part collection] created by the 2015 UCLA iGEM Team. For more information on ICA, please visit the ICA protocols [http://2015.igem.org/Team:UCLA/Notebook/Spider_Silk_Genetics/Protocols page] and the collection parts pages.

This part was sequence verified using registry standard VF2 and VR primers.

Source

This is derived from the Major Ampullate Spidroin (MaSp) protein consensus sequences found in the spider N. Clavipes. (Hinman and Lewis, 1992 and Xu and Lewis, 1990) The MaSp comes in two forms, MaSp1 and 2, both of which confer different properties on the final fiber. MaSp1 confers strength, and MaSp2 confers elasticity. We back-translated the protein to derive a suitable coding sequence. We optimized the coding sequence for a reduced GC content (~59% GC), rather than for the organism (~75% GC). This was done to allow for normal PCR without having excessive GC pairs.

References

Briggs A., Rios X., Chari R., Luhan Y., Zhang F., Mali P., and Church G. Iterative capped assembly: rapid and scalable synthesis of repeat-module DNA such as TAL effectors from individual monomers. Nucleic Acids Research. 2012;40(15): e117

Hinman, M.B., Lewis, R. Isolation of a clone encoding a second dragline silk fibroin. Nephila clavipes dragline silk is a two-protein fiber. J. Biol. Chem. 1992;267: 19320–19324.

Tokareva O., Michalczechen-Lacerda V., Rech E., and Kaplan D. Recombinant DNA production of spider silk proteins. Microbial Biotechnology. 2013;6(6): 651-663

Xu, M., Lewis, R.V. Structure of a protein superfiber: spider dragline silk. PNAS;1990:87: 7120.