Re: [DIYbio] Re: Seaweed shortage will impact agar prices

18kbp is nothing. Everyone should stop using biobricks... Overlap pcr or in-vitro cloning should be the main teaching and building tool for iGEM, but that is going off topic...and also my oppinion.

Sebastian S. Cocioba

CEO & Founder

New York Botanics, LLC

Blog: ATinyGreenCell.com

On Dec 14, 2015, at 5:51 AM, Yuriy <yuriy...@gmail.com> wrote:

Not from my searches. How do you plan to purify it from a higher plant?

 

A search of "alginate synthesis" site:igem.org shows up as this paper: "Genome-Scale Metabolic Network Analysis of the Opportunistic Pathogen Pseudomonas aeruginosa PAO1", that cites, ref 46; http://www.ncbi.nlm.nih.gov/pubmed/15813726

 

I could have saved a ton of time with a simple "bacterial alginate synthesis." Turns out it is in biofilm production for pathogenic strains of P. aregunosa. There is this runaway process of overproduction of the exopolysaccharide alginate.

http://www.ncbi.nlm.nih.gov/pmc/articles/PMC2435354/ well characterized. Also bacterial.

See table 1. Have fun 18+ kbp. how much do you like to BioBrick? 

How ironic. if it microbiology may now be the source of the very thing it was cultured on all along.

Trehalose is used in DNA and Protein preservation. Maybe there's an avenue to explore for a gellating agent. 

why not Nannochloropsis for a candidate species? "two birds with one stone" sort of thing. 

On Saturday, December 12, 2015 at 11:41:06 AM UTC-5, Sebastian wrote:

Now thats a fun topic! Anyone know what the genes encoding the synthesis of agarose are? Think simple cytosolic expression would gunk up the internals of the cell? If we ship it to the vacuole of a higher plant, would it interfere? If there is only one or two commercial sources for the stuff (red algae) then why not move it to chlorella? Would it really be more cost effective? Has anyone done this in bacteria (iGEM)? If there is a POC in E. Coli, you could horizontally transfer it to algal chloroplasts...issue would be export out to avoid occluding the thylakoids and shut down the photosystems...which are kinda important.

Anyone know of other gel biopolymers that are clear, inert, and cheap? Would at least be an interesting thought experiment if we could design a circuit for agar expression via nuclear transformation (agro), or just one for bacteria like cyano. Thoughts? 

Sebastian S. Cocioba

CEO & Founder

New York Botanics, LLC

Blog: ATinyGreenCell.com

On Dec 12, 2015, at 10:57 AM, Brian Degger <brian....@gmail.com> wrote:

Simon, 

that was too obvious...someone probably has that in a draft somewhere. 

cheers

Brian

On Sat, Dec 12, 2015 at 3:41 PM, Simon Quellen Field <sfi...@scitoys.com> wrote:

In this group I would expect someone to mention placing genes from seaweed into a similar alga that was easier to culture, and perhaps amplifying the agarose output. Did I miss that part of the thread?

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On Sat, Dec 12, 2015 at 2:48 AM, Cathal Garvey <cathal...@cathalgarvey.me> wrote:

Worth adding here btw that I experimented ages back with isolating
agarose from agar. The outcome was "possible, but really awkward and
ultimately not even close to price-competitiveness as buying agarose".
Put another way, agarose seems expensive but is actually sold for
significantly cheaper than it'd take to make your own.

However, when all you care about is making research-grade agar, things
might be a bit rosier. For example, it won't get you anywhere near
"agarose", but you can improve the grade of agar by simply reducing the
agaropectin content using pectinase. I didn't find any useful sources on
how to set up a pectinase reaction with agar, so "here be dragons", but
if it were possible to simply hydrate cheap, crappy-grade food agar and
add brewer's pectinase after adjusting pH with vinegar and carbonates,
then leaving for a few days..

I recommend making up the "buffer" first with enzyme at your favourite
pH and then adding the agar, so the hydration of the agar beads can pull
in the dissolved/suspended enzyme. This will also give you a chance to
decant any contaminants out of your "buffer" so you don't get
contaminating nutrients into your agar. Though, with food agar, it's
probably nutritionally "dirty" enough already.

Pectinase is used to clarify fruit mashes and wines in brewing, so
perhaps having some "cloudy" fruit juice next to your agar, and
comparing progress on clarification as a proxy for agaropectin
degredation..though I expect the latter reaction will take longer
because agaropectin isn't "free pectin" and will be bound within agar
grains.

You might then have to wash out the degraded pectins, and there are
plenty of avenues to do that. You could make a thin gel and press out
the fluid to get agar flakes, or you could simply make sure your agar
grind is quite fine before treating it, so the pectins can dissolve in a
straightforward rinse..

On Thu, 2015-12-10 at 16:06 -0500, Sebastian S Cocioba wrote:
> Gelrite (gelzan) is nice for its clarity. A little soft and more
> expensive than agar agar. It is also fairly sensitive to pH and is
> basically slush around 5.0-5.3 (I learned that the hard way). It also
> doesn't like high phosphates like that in M9. Its good in some cases
> for plant tissue culture, bad for pretty much everything else. There
> was this pallet sold on eBay for $80/kg of gelrite and I gobbled up
> four tubs. Street price is like $160 and now this little publicity
> stunt will most likely artificially drive prices up before any real
> shortage occurs.
>
>
>  You could use any agar if you just test for gel strength. I made a
> little jig a while back to measure gel strength relative to 1000g/cm3
> agar using a rubber band, pen, and some popsicle sticks. It just
> measures how high a pen attached perpendicularly to a flat 1cm^2 can
> rise on a scale when force from rubber band acts on it. Basically
> force till smush. Calibrate it with a weight for analytical balance
> calibration and derive force. You can also use muscle memory if you
> culture a lot and know when its right. I can tell if I messed up the
> pH in my MS based on the jiggle in the media bottle and the way the
> seeds embed when dropped. I know thats not very scientific but agar
> varies so much batch to batch and strain to strain (algal species like
> carigeean) that sometimes you need a more instinctive understanding.
> This of course is not too necessary for routine bacteria. If you flick
> an upside down plate and it vibrates a bit, its dense enough. For
> plants its a whole different story. Density impacts root formation
> greatly and some just wont root in soft or too hard agar. Interesting
> things you learn when you do this for too long...
>
> Sebastian S. Cocioba
> CEO & Founder
> New York Botanics, LLC
> Blog: ATinyGreenCell.com
>
>
>
> On Dec 10, 2015, at 12:48 PM, Dakota Hamill <dko...@gmail.com> wrote:
>
>
> > Time to stock up and re-sell for a profit.
> >
> >
> > I've heard GelRight is a good alternative.  Supposedly, it's even
> > more "inert" than agar.
> >
> >
> > http://www.sigmaaldrich.com/catalog/product/sigma/g1910?lang=en&region=US&gclid=COTBru_s0ckCFVQYHwodmCsLTg
> >
> >
> >
> >
> >
> >
> > On Thu, Dec 10, 2015 at 12:46 PM, Yuriy <yuriy...@gmail.com>
> > wrote:
> >         Time to look for alternatives.
> >
> >         On Thursday, December 10, 2015 at 9:49:46 AM UTC-5, Koeng
> >         wrote:
> >                 http://www.nature.com/news/lab-staple-agar-hit-by-seaweed-shortage-1.18970
> >
> >
> >                 -Koeng
> >         --
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