Part:BBa_K1692002
codon optimized FDC with T7 promoter and Flag Tag
Overview
Ferulic acid decarboxylase (FDC) catalyzes the conversion of trans-cinnamic acid to styrene. We codon-optimized S. cerevisiae’s FDC gene for expression in E. coli. This construct includes a T7 inducible promoter, a ribosome binding site, and a FLAG-tag peptide sequence for easy and efficient protein purification. We have sequenced our construct and verified that all these components are indeed present. We were able to successfully clone our construct into E. coli and induce expression with isopropyl β-D-1-thiogalactopyranoside (IPTG). A sodium dodecyl sulfate polyacrylamide gel electrophoresis confirmed that our FLAG-tag extraction selectively purified the FDC enzyme.
Experiments and Results
Reference
[1] Mckenna, Rebekah, Luis Moya, Matthew Mcdaniel, and David R. Nielsen. "Comparing in Situ Removal Strategies for Improving Styrene Bioproduction." Bioprocess Biosyst Eng Bioprocess and Biosystems Engineering (2014): 165-74. Print.
[2] White, Mark D., Karl A. P. Payne, Karl Fisher, Stephen A. Marshall, David Parker, Nicholas J. W. Rattray, Drupad K. Trivedi, Royston Goodacre, Stephen E. J. Rigby, Nigel S. Scrutton, Sam Hay, and David Leys. "UbiX Is a Flavin Prenyltransferase Required for Bacterial Ubiquinone Biosynthesis." Nature (2015): 502-06. Print.
Sequence and Features
- 10COMPATIBLE WITH RFC[10]
- 12COMPATIBLE WITH RFC[12]
- 21INCOMPATIBLE WITH RFC[21]Illegal BglII site found at 233
- 23COMPATIBLE WITH RFC[23]
- 25COMPATIBLE WITH RFC[25]
- 1000COMPATIBLE WITH RFC[1000]
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