Is it possible for a Homeworker to create a thermosensitive yoghurt?

[Lukas]

Hey Guys

I want to create a thermosensitive yoghurt which reacts with bioluminescence under heat. I have the model of Meredith Patterson who has created a test for Melamin. I want to create the pudding for a school Project.

Lg Lukas

BTW if somebody has time : can you list the equipment for the experiment?

[Koeng]

In my experience, visible lux takes some time to produce. Perhaps just bioluminesence would be better, heat activation brings in a lot of complications

Also, it takes quite a while to create yogurt strains (that I've worked with). I worked with one strain, S thermophilus, and it was a bit finicky with growth. Still, there's nothing stopping you from doing it.

If you have a bit bigger budget, perhaps pay someone to do the cloning of the lux operon for you. I have one lux (not tested with visible light production) cassette for Bacillus if needed, it's quite similar to S thermophilus and might be able to work.  You'll be able to work quicker but perhaps not learn as much about cloning if you just get a company to clone the DNA, you can just work directly with yogurt. You also wont get very much variation, so troubleshooting would be more difficult

Where are you located? Perhaps a local biohacker lab can help you out with you idea

-Koeng

[Mega [Andreas Stuermer]]

I also got pVIB, but the bacillus stuff will work better in streptococcus and lactobacillus (gram-)

Lux luciferase is denatured above 30°C, so it stops glowing. There's also a vibrio YFP which shifts the emitted colour to yellow. Though this YFP only works below 20°C, else you get the usual blue light

[Jonathan Cline]

On Thursday, January 22, 2015 at 2:46:49 AM UTC-8, Lukas wrote:

It will take you years to develop this, if it ever works at all, though by the time you are done, you might have a Ph D.  The equipment list is partially in the DIYbio FAQ  http://openwetware.org/wiki/DIYbio/FAQ

You probably want to look into Genomikon.


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[Cathal Garvey]

Actually, I think this is a *much* easier proposition than the melamine
sensor (which never happened AFAIK). You can couple a gene system to
heat-shock promoters, and you're mostly there.

The complication is that the luciferin molecules are, I think, pretty
heat-labile. So, it might not work simply because the luciferin is being
pasteurised prior to oxidation by luciferase.

Also, some "heat" shock systems are actually just "shock" systems, and
may also activate under colder-than-ideal growth temperatures, so you
might find that what you really have is "glows when not at exactly 37C".

Covering for leaky expression would be important, but not that
important. As long as the luciferin/ase production is at the end of the
regulatory cascade and it scales nicely to input, some leaky expression
will be pretty invisible.

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[Jonathan Cline]

As the original question was:
Q: "Is it possible for a Homeworker to create a thermosensitive yoghurt?"

My answer is: "No."
Oh, even if it's "just" for a school pudding project, it's still: "No."

Genomikon does have glowing smelly things which are growable for a
school pudding project for a Homeworker, and not thermosensitive unless
one counts 'cell death' as being sensitive which is unlikely.

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[Nathan McCorkle]

On Wed, Jan 28, 2015 at 5:55 AM, Cathal Garvey
<cathal...@cathalgarvey.me> wrote:
> Actually, I think this is a *much* easier proposition than the melamine
> sensor (which never happened AFAIK). You can couple a gene system to
> heat-shock promoters, and you're mostly there.
>
> The complication is that the luciferin molecules are, I think, pretty
> heat-labile. So, it might not work simply because the luciferin is being
> pasteurised prior to oxidation by luciferase.

Couldn't you heat-shock to induce enzyme production, then spike with
luciferin after the culture cools?

[Mega [Andreas Stuermer]]

Why do you want to add external luciferin? In bacteria, the vibrio fischeri lux system works pretty well and does not require external substrates

[Filip Hasecke]

Hi Lukas,

it might actually be possible. I found a fluorescent protein that experiences an absorbance shift in a range between 4°C and 50°C.

Take a look here: http://www.nature.com/srep/2012/120829/srep00608/full/srep00608.html

The protein that is used in that study is called WasCFP (derived from mCerulean by mutagenesis). You might be able to transform your bacteria with a plasmid carrying a gene for that protein. Then you might be able to produce a heat sensitive yoghurt. ;)

I did not take a very close look into it yet, but that might be the right direction.

Cheers

Filip

2015-01-28 21:42 GMT+01:00 Mega [Andreas Stuermer] <masters...@gmail.com>:

Why do you want to add external luciferin? In bacteria, the vibrio fischeri lux system works pretty well and does not require external substrates

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[Nathan McCorkle]

On Wed, Jan 28, 2015 at 12:42 PM, Mega [Andreas Stuermer]
<masters...@gmail.com> wrote:
> Why do you want to add external luciferin?

I guess I didn't see/remember any discussion about adding/producing
this, so just assumed that was the idea.

> In bacteria, the vibrio fischeri
> lux system works pretty well and does not require external substrates

So can't this just be induced with heat-shock too? Are there any
timers that are easily usable? Something that is known to be an
autoinducer... like the quorum sensing systems:

Autoinducers Act as Biological Timers in Vibrio harveyi
http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0048310

[Mega [Andreas Stuermer]]

You could induce luxI with a thermal promoter. LuxI produces the homoserine lacton that activates luxICDABEG expression (actually it doesnt activate but induce it 1000 fold)