[DIYbio] Re: [diybio-eu] Re: IndieBB Crowdfunding Campaign: Help me make a great beginner's kit for DIYbio/synbio!

I guess you'd only need this ability if you were in some trade-control
zone, where getting a strain would be hard/impossible to do. Not sure
if many countries like that exist, but I can think of at least one. Or
maybe in an area where postal delivery from a kind friend is
hard/expensive/unavailable... I can think of more than one country
like this, or remote/undeveloped areas inside countries.

On Wed, Jan 29, 2014 at 1:44 PM, Cathal (Tablet)
<cathalgarvey@cathalgarvey.me> wrote:
> You see, with the right signalling it'd be no problem. But, this is to be a
> 'hackable' plasmid, meaning someone might want to build a project on it that
> requires signals like temperature, tetracycline, etcetera. If I were to use
> those signals as a kill-switch for the plasmid, it'd make the plasmid
> incompatible with projects using those signals.
>
> Synthetic Biology is already short on useful inputs; there aren't that many
> distinct, independent signals you can use at the same time for complex
> projects. Better if I use *none* in the plasmid, and leave them all open for
> people to use.
>
> By and large, plasmid-curing isn't a feature most people want in a cloning
> or engineering plasmid. If you do want it, hack it in; create an inducible
> system with antisense RNA against the toxins and transporters but not the
> immunity protein, and to a lesser extent against the rep protein. So, the
> culture will (simultaneously) stop making toxin, survive for a while, and
> eventually lose the plasmid due to low rep protein levels. But, a killswitch
> like this isn't a feature I'd bother putting into the plasmid on day 1.
>
>
> Andreas Stuermer <masterstorm123@gmail.com> wrote:
>>
>> You would need a silencer RNA that inhibits the toxin gene at 30°C and a
>> replication protein that loses its function at 30°C. That would work well.
>>
>>
>> On Wed, Jan 29, 2014 at 8:23 PM, Andreas Stuermer
>> <masterstorm123@gmail.com> wrote:
>>>
>>> The replication protein can be repressed by tetracyclin? I think the
>>> bacteria can sense tetracyclin long before they actually get strongly
>>> inhibited?
>>>
>>> Or this temperature gene? Above 30°C the replication protein is no longer
>>> produced and thus the plasmid is lost without selection. Actaully - there is
>>> selection. Except if you dilute them very very much. Hm... the toxin will
>>> have a longer shelf-live than the antitoxin - thus if the bacterium loses
>>> the plasmid it gets killed by the remining toxin. That's the sense of it
>>> actually...
>>>
>>>
>>> On Wed, Jan 29, 2014 at 8:12 PM, Nathan McCorkle <nmz787@gmail.com>
>>> wrote:
>>>>
>>>> So I started wondering last night, do you have any ideas on how to
>>>> cure a strain of such a plasmid? I could imagine someone running out
>>>> of wild/plasmidless bacteria, or maybe their lab getting contaminated
>>>> with the plasmid and thus causing them to have no more plasmidless
>>>> E.coli (plasmidless in the sense of your engineered plasmid).
>>>>
>>>> With antibiotic resistance plasmids, you can usually 'cure' them of
>>>> the plasmid by growing them without pressure (no antibiotic added) for
>>>> several generations. Is there any way to 'cure' these bugs with your
>>>> plasmid? Some 'curative' plasmid that interferes with transcription of
>>>> the colicin? Seems like to then get rid of the curative plasmid, it
>>>> would have to be controlled externally (i.e. with antibiotics).
>>>>
>>>> Thoughts?
>>>>
>>>> On Wed, Jan 22, 2014 at 12:01 PM, Cathal Garvey
>>>> <cathalgarvey@cathalgarvey.me> wrote:
>>>> > Hey all,
>>>> > As anyone on this list for more than a month can attest, our most
>>>> > common
>>>> > FAQ here is "How do I get started?". More often than not, it's more
>>>> > specifically "How do I get started in synthetic biology?", and our
>>>> > answers can get a bit woolly.
>>>> >
>>>> > The truth is that our options for beginners all suck. Most suppliers
>>>> > are
>>>> > hostile towards independent buyers, so we tell new people to buy the
>>>> > "refill kit" from Carolina and pretend to be a school, or we mail
>>>> > plasmids of dubious provenance to one another. While that's good for
>>>> > community spirit, it says a lot that we celebrate knowing the
>>>> > approximate sequence of one of them at last.
>>>> >
>>>> > Finally, we've talked a lot about the issue of getting, using and
>>>> > disposing of antibiotics for this purpose a lot, and we've talked more
>>>> > and more recently about removing the need for antibiotics altogether.
>>>> >
>>>> > That's what I aim to do, and I'd really appreciate your help and
>>>> > support
>>>> > making it happen. Here's my IndieGoGo campaign, freshly pressed:
>>>> > http://www.indiegogo.com/projects/indiebb-your-first-gmo
>>>> >
>>>> > The kit is designed for beginners, and for teachers and education
>>>> > groups, to introduce people to the methods of E.coli engineering. It's
>>>> > also designed to be hackable and to be fairly licensed in an Open
>>>> > Source
>>>> > way that preserves the user's freedoms going forward. It's fairly
>>>> > priced, and designed by and for DIYbioers. It'll be fluorescent, and
>>>> > it
>>>> > won't need antibiotics.
>>>> >
>>>> > If you're interested, please help me out and support the IndieGoGo
>>>> > campaign. It's an all-or-nothing campaign, so if I don't hit the goal,
>>>> > nothing is raised. If you know a bio/hacker, educator or student who'd
>>>> > be interested, please let them know, too. And if you're on Twitter or
>>>> > (ugh) Facebook, a shoutout to let others know would be really, really
>>>> > appreciated. :)
>>>> >
>>>> > For the purposes of fundraising and separating my usual noise from the
>>>> > campaign, I've started a new twitter account for this, too:
>>>> > @IndieBBDNA
>>>> >
>>>> > Gratefully yours, and looking forward to collaborating on this and
>>>> > making it real!
>>>> > Cathal
>>>> > PS: As I know too well, nothing can be guaranteed to work in Synbio at
>>>> > this point, so bear that in mind. However, this is the most
>>>> > conservative
>>>> > design I've yet embarked on, and I'm confident it will work. So, bear
>>>> > that in mind too. :)
>>>>
>>>>
>>>>
>>>> --
>>>> -Nathan
>>>>
>>>> ---------------------------------------------------------------------
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>>>> For additional commands, e-mail: diybio-eu-help@diybio.eu
>>>>
>>>
>>
>
> --
> Sent from my Android device with K-9 Mail. Please excuse my brevity.



--
-Nathan

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