Difference between revisions of "Part:BBa K2916007:Design"
 
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===Design Notes===
 
===Design Notes===
  
−
The sequence was extracted from the pET21a-GlyRS-His plasmid and expressed in the BL21 (DE3) E.coli strain.  
+
The sequence was extracted from the pQE30-His-PheRS plasmid and expressed in the M15 E.coli strain.
 +
 
 +
Codon optimization : 810 from G to C
 +
 
 +
Codon optimization : 2151 from G to C
 +
 
 +
Second stop codon was added in compliance with iGEM standards.
  
 
===Source===
 
===Source===
  
−
http://www.addgene.org/124110/
+
Sebastian Maerkl Lab at EPFL, Switzerland.
 +
 
 +
http://www.addgene.org/124116/
  
 
===References===
 
===References===
 +
 +
<i>
 +
McClain, William H. “Rules That Govern TRNA Identity in Protein Synthesis.” Journal of Molecular Biology, vol. 234, no. 2, Nov. 1993, pp. 257–80. DOI.org (Crossref), doi:10.1006/jmbi.1993.1582.
  
 
Cell-free translation reconstituted with purified components. Shimizu Y, Inoue A, Tomari Y, Suzuki T, Yokogawa T, Nishikawa K, Ueda T. Nat Biotechnol. 2001 Aug;19(8):751-5. doi: 10.1038/90802. 10.1038/90802 PubMed 11479568
 
Cell-free translation reconstituted with purified components. Shimizu Y, Inoue A, Tomari Y, Suzuki T, Yokogawa T, Nishikawa K, Ueda T. Nat Biotechnol. 2001 Aug;19(8):751-5. doi: 10.1038/90802. 10.1038/90802 PubMed 11479568
 +
 +
Lavickova, Barbora, and Sebastian J. Maerkl. A Simple, Robust, and Low-Cost Method to Produce the PURE Cell - Free System. preprint, Synthetic Biology, 18 Sept. 2018. DOI.org (Crossref), doi:10.1101/420570.</i>

Latest revision as of 02:02, 22 October 2019


PheRS protein equipped with a 6x HIS affinity tag


Assembly Compatibility:
  • 10
    COMPATIBLE WITH RFC[10]
  • 12
    COMPATIBLE WITH RFC[12]
  • 21
    INCOMPATIBLE WITH RFC[21]
    Illegal BglII site found at 2424
    Illegal BamHI site found at 31
    Illegal BamHI site found at 1772
  • 23
    COMPATIBLE WITH RFC[23]
  • 25
    INCOMPATIBLE WITH RFC[25]
    Illegal NgoMIV site found at 1162
    Illegal AgeI site found at 1155
    Illegal AgeI site found at 2077
    Illegal AgeI site found at 2929
  • 1000
    INCOMPATIBLE WITH RFC[1000]
    Illegal BsaI.rc site found at 2875
    Illegal SapI.rc site found at 198
    Illegal SapI.rc site found at 2851


Design Notes

The sequence was extracted from the pQE30-His-PheRS plasmid and expressed in the M15 E.coli strain.

Codon optimization : 810 from G to C

Codon optimization : 2151 from G to C

Second stop codon was added in compliance with iGEM standards.

Source

Sebastian Maerkl Lab at EPFL, Switzerland.

http://www.addgene.org/124116/

References

McClain, William H. “Rules That Govern TRNA Identity in Protein Synthesis.” Journal of Molecular Biology, vol. 234, no. 2, Nov. 1993, pp. 257–80. DOI.org (Crossref), doi:10.1006/jmbi.1993.1582.

Cell-free translation reconstituted with purified components. Shimizu Y, Inoue A, Tomari Y, Suzuki T, Yokogawa T, Nishikawa K, Ueda T. Nat Biotechnol. 2001 Aug;19(8):751-5. doi: 10.1038/90802. 10.1038/90802 PubMed 11479568

Lavickova, Barbora, and Sebastian J. Maerkl. A Simple, Robust, and Low-Cost Method to Produce the PURE Cell - Free System. preprint, Synthetic Biology, 18 Sept. 2018. DOI.org (Crossref), doi:10.1101/420570.