Re: [DIYbio] Vibrio fischeri isolation

Ah, ok... So the substrate that lux CDE uses, right? So adding those genes which make the precursor will enhance luminescence much more than gene copy?

Well, I hope it's V.fischeri, because the operon is very compact. So one PCR with two primers can ghet you all the lux genes needed.

The structure of P.Phosphoreum is more seperated. IIRC, the flavin reductase gene is not even in the operon -> 2 PCRs needed



On Fri, Nov 30, 2012 at 3:59 PM, Cathal Garvey <cathalgarvey@gmail.com> wrote:
Yea, copy number of the DNA isn't the limiting factor in
bioluminescence: quantity of inputs is much more limiting.
You can get great luminescence from natural Vibrio species, and they can
be easier to grow in pure culture if you isolate Vibrio phosphoreum
rather than Vibrio fischeri (if it's bright blue and can grow at 4C,
odds are it's phosphoreum).

Sadly, my cultures of phosphoreum don't seem to have survived long-term
freezer storage, but they're satisfying to re-isolate when lost!

Unfortunately, they smell pretty bad during growth.. :P
According to some guides I've seen, you can maximise their brightness by
feeding them glycerol rather than glucose/sucrose. This might be due to
whether or not they produce acid from glycerol or not; acid would
probably screw up bioluminescence.

On 29/11/12 17:04, Mega wrote:
> Hi, two days ago I bought a piece of Salmon, cut away the skin. I placed
> the skin on a plate and put some salty water (30g/L) on the plate, yet
> leaving the salmon-skin unwetted.
>
> Left it in our basement (some 10-15°C maybe).
>
>
>
> As I had a look today, it was amazing. They are very bright. (They are
> nearly as bright as the pVIB E Coli were, althought the Colis had the
> operon ~20 times (medium copy based on pBR322), while V. Fischeri has
> just a single copy of the operon).
>
> You just walk into the room, switch of the light and it glows amazingly
> (i.e. visibly without eye adaption).
>
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