Lux operon

From 2006.igem.org

This is a page about the Lux operon (CiteULike) and the technicalities of making it into a Part in the Registry.

Background

So the main task, as I understand it, is this (and I might have some of the specifics a bit wrong, so take this with a grain of salt): we want to make a new Part that contains the Lux operon (for clarity I am going to capitalize "Part" when I use it in the sense of a genetic Part from the registry). The physical manifestation of a Part is a special plasmid called a "biobrick". There are variety of biobricks, but all have fundamentally the same design: all contain an antibiotic resistance gene (a selector), an origin of replication, perhaps some other things, and the actual Part + "biobrick ends." The biobrick ends are a specific sequence of restriction sites which bracket the Part in a standard way, so that this section of the plasmid is always organized in the following manner: EcoRI - XbaI - INSERT - SpeI - PstI, where INSERT designates the actual Part ( i.e. the Lux operon). It is essential that these four particular restriction sites occur only once, in the given order, in the plasmid, for their standard organization and single occurance in each biobrick enables their modularity. Hence the INSERT - the Part - cannot contain any of the four restriction sites. I haven't checked if the Lux operon contains any of them, but it's pretty big, so chances are it does, and thus our primary challenge will be rengineering the operon to lack these sites (via the introduction of silent point mutations).

Questions

• Is there more than one version of the Lux operon?
  • If so, which one do we use?
  • How do we decide which will be most likely to work?
  • What do they all have in common and what makes them different? The inducer?
• Where do we get it?
• Into which biobrick chassis should we insert it?

Progress

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