Team:Copenhagen/Parts

From 2012.igem.org

Aspects

An important aspect of the iGEM competition is the use and creation of standard biological parts. Each team will make new parts during iGEM and will place them in the Registry of Standard Biological Parts. The iGEM software provides an easy way to present the parts your team has created . The "groupparts" tag will generate a table with all of the parts that your team adds to your team sandbox. Note that if you want to document a part you need to document it on the Registry, not on your team wiki. Remember that the goal of proper part documentation is to describe and define a part such that it can be used without a need to refer to the primary literature. The next iGEM team should be able to read your documentation and be able to use the part successfully. Also, you should provide proper references to acknowledge previous authors and to provide for users who wish to know more.

Parts submitted

BBa_K770000 Composite part - ProC-YF1 – in pSB1C3 backbone plasmid
This biobrick consists of two parts:
  • ProC (10 bp): A constitutively active promoter active in cyanobacteria.
  • YF1 (1131 bp): A chimeric protein part of a synthetic light-inducible regulatory system. YF1 is a kinase which autophosphorylates and phosphorylates its substrate, FixJ. The activity of YF1 is inhibited by light.

Design Notes:
Partial assembly of the planned complete construct of the 2012 iGEM Team Copenhagen: CyanoDelux. The part has been assembled using USER-cloning. This leaves small scars comprised of USER overhangs. These should not interfere with the function of the part.


BBa_K770001 Reporter gene: LuxCDABE gene cassette - In pSB1C3 backbone plasmid
The LuxCDABE gene cassette (5798 bp) comprises 5 genes necessary for emission of light with a wavelength of approximately 490 nm.
  • LuxA and LuxB encode two polypeptides of 40 kDa and 37 kDa, respectively, which together make up the complete bacterial luciferase heterodimer. Luciferase catalyzes a light-producing reaction in which the substrates fatty aldehydes, O2 and FMNH2, are degraded into fatty acid, H2O and FMN.
  • Together the Lux C, D and E encode a Fatty Acid Reductase Enzyme complex which converts acyl-doners to fatty acid, then fatty aldehydes.
  • Lux D: encodes a transferase which transfers acyl-donors from the fatty acid synthetic pathway, transforming acyl-doners to a fatty acid.
  • Lux E: encodes a synthetase which, using ATP, converts the fatty acid to an acyl-AMP intermediate,
  • Lux C: encodes reductase, which consumes NADPH to release fatty aldehyde and NAD+.

Design Notes:
The part was assembled with USER-cloning (Read more), incorporating a complete LuxCDABE cassette into a pSB1C3 plasmid. The procedure has left small scars in the form of USER overhangs, which should not affect activity or regulation of the gene cassette. The LuxCDABE gene contained an illegal Xbal-site which was subsequently removed using PCR site-directed mutagenesis.
The sequences of the scars are given below.
AGTGCGAT
ACTTGCGT

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