|Project Collaborators: Open Bioeconomy Lab (University of Cambridge)|
Individual Contributors: Jenny Molloy (University of Cambridge), Chiara Gandini (University of Cambridge)
Supported by: Shuttleworth Foundation
Scott Pownall helped in retrieving the sequences of luciferase genes.
This spreadsheet documents Reporter genes sequences whereby the US patent covering the enzyme sequence itself has expired.
The uses of that enzyme for molecular biology and biotechnology protocols and applications may still be covered by patents in various countries and we are compiling a list of these separately.
|Key to column headings|
|Reporters (name)||Name or commonly used description of the Reporters. Equivalent trademarked names are provided for reference where applicable.|
|Features and Applications||Features of the polymerase and its applications in molecular biology and biotechnology|
|Patent Expired||Whether the patent covering the enzyme or its sequence has expired.|
|US Filing/Application Date||Priority date for the first US patent covering the enzyme. This is the date at which protection for the invention begins.|
|US Expiration Date||In the United States, for utility patents filed on or after June 8, 1995, the term of the patent is 20 years from the earliest filing date of the application on which the patent was granted and any prior U.S. or Patent Cooperation Treaty (PCT) applications from which the patent claims priority (excluding provisional applications). For patents filed prior to June 8, 1995, the term of patent is either 20 years from the earliest filing date as above or 17 years from the issue date, whichever is longer. We have used this formula except in cases where i) an extension to term or a termination has been specified in the patent; ii) the patent is listed as expired due to lack of payment of fees.|
|Original Paper||The paper where the enzyme is initially described, in some cases this is the original discovery and in some cases the first characterisation of the recombinant form or where possible both cases are provided.|
|Original Patent Family||The first patent family covering the enzyme, which may not include all subsequent modifications.|
|Notes||Any additional notes relevant to the enzyme or patents|
|Max excitation (nm)||The wavelength (nm) at which the fluorophore reacts the most|
|Max emission (nm)||The main wavelength (nm) at which the fluorophore will emit light, following excitation|
|Quantum yield||Is the ratio of photons emittet to photons absorbed. A high QY is desiderable.|
|Molar Extinction||Defines how strongly a fluorophore attenuates light of a given wavelength (high greater amount of light being absorbed)|
|Original DNA sequence||DNA sequence retrived from the original organism|
DNA Sequence optimized and domesticated
|DNA sequence as present in the collection. The sequences are codon optimized to be expressed in E. coli (see "Notes") and domesticated to avoid restriction sites used in Biobrick and GoldenGate/Loop/MoClo cloning strategies (BamHI, BglII, EcoRI, NheI, NotI, NruI, PstI, SpeI, XbaI, XhoI, BbsI, BsaI, BsmbI, SapI)|
|Protein Sequence||Protein sequence and data from the Uniprot database|
|Twist synthesis score||Is the score that the sequence will have if submitted to Twist for synthesis. It could be either Standard, Difficult, Impossible or Too Short (when possible, sequences were adjusted to give "standard" score e.g. strings of 9 As (coding for 3 Ks) make sequences impossible to synthesize, these were broke changing the middle codon AAA->AAG; when "too short" the sequence requires an addition of dummy DNA stretches to be synthesized.|
Original papers from where most of the fluo- and chromoproteins come from:
Liljeruhm, Josefine, et al. "Engineering a palette of eukaryotic chromoproteins for bacterial synthetic biology." Journal of biological engineering 12.1 (2018): 8.
Alieva, Naila O., et al. "Diversity and evolution of coral fluorescent proteins." PLoS one 3.7 (2008): e2680. https://journals.plos.org/plosone/article?id=10.1371/journal.pone.0002680
|NOTES on the Reporter collection|
|1) although some proteins might look ideantical in amino acids sequence, this does not mean that their properties are the same: a 98% identity can lead to a substantial differences in quality of light emission. 2)all the partially characterized proteins (e.g. from igem) have been added only if off-patent. If a protein not part of the iGEM collection was found and this protein was very similar or identical in spectral characteristics to a proteins of iGEM collections but showed substantial improvements in QY or ME values, that protein was added in the collection 3) a protein was excluded from the list if found identical or very similar for all the parameters max excitation/max emission/QY/ME to other proteins already present in the list. 4) some proteins have been already partially characterized (e.g. in iGEM competitions) and have been included in the list even if their characteristics are not as good as other similar proteins already present in the list|