Seaweed shortage will impact agar prices
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Koeng
Dec 10, 2015, 9:49:46 AM12/10/15
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Yuriy
Dec 10, 2015, 12:46:57 PM12/10/15
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Time to look for alternatives.
On Thursday, December 10, 2015 at 9:49:46 AM UTC-5, Koeng wrote:
On Thursday, December 10, 2015 at 9:49:46 AM UTC-5, Koeng wrote:
Dakota Hamill
Dec 10, 2015, 12:48:31 PM12/10/15
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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.
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Cathal (Phone)
Dec 10, 2015, 3:17:49 PM12/10/15
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I concur: synthetic alternatives are overdue, especially if concerns about ROS generation when autoclaving agar (and the possible.link to unculturable bacteria) are true.
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Bryan Jones
Dec 10, 2015, 3:33:25 PM12/10/15
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You could always go back to culturing microbes on potato slices.
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Sebastian S Cocioba
Dec 10, 2015, 4:06:35 PM12/10/15
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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
Sebastian S. Cocioba
CEO & Founder
New York Botanics, LLC
Blog: ATinyGreenCell.com
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Cathal Garvey
Dec 12, 2015, 5:48:36 AM12/12/15
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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..
> > +unsub...@googlegroups.com.
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..
Gordana Ostojic
Dec 12, 2015, 10:30:32 AM12/12/15
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capillary electrophoresis?
Simon Quellen Field
Dec 12, 2015, 10:42:22 AM12/12/15
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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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Brian Degger
Dec 12, 2015, 10:58:24 AM12/12/15
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Simon,
that was too obvious...someone probably has that in a draft somewhere.
cheers
Brian
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Brian Degger
Brian Degger
twitter: @drbrian
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Sebastian S Cocioba
Dec 12, 2015, 11:41:06 AM12/12/15
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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?
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Yuriy
Dec 14, 2015, 5:51:30 AM12/14/15
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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.
why not Nannochloropsis for a candidate species? "two birds with one stone" sort of thing.
Sebastian S Cocioba
Dec 14, 2015, 12:21:59 PM12/14/15
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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.
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Koeng
Dec 14, 2015, 7:11:57 PM12/14/15
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I agree, making your own gibson/ SLiCE mix has gotten cheap and efficicent enough to the point of where there really isn't a reason to use Biobricks anymore.
Although my opinion is that iGem should begin using a standard system of GoldenGate so that they can retain some of their original goal of modularity and ability to ship a large registry of standard parts.
-Koeng
Sebastian S Cocioba
Dec 14, 2015, 7:42:44 PM12/14/15
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I've found the BsaI and Btgz...whatever sites in most of my wild type isolated cds. Is it wise to promote a system that is so dependent on synthesis? Too soon for too high a cost? Sorry to go wildly off topic...
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Xabier Vázquez Campos
Dec 14, 2015, 7:45:08 PM12/14/15
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Sebastian, Gelrite (gellan gum) needs divalent cations to gel in normal conditions. I've use it for culturing fungi even at pH 1.0. Although the concentration needed may vary. Most media contain absurdly high phosphate concentrations and some studies have shown that many, if not most, bugs can grow even better with much lower levels or even just traces of phosphates in the media
Cathal, the ROS generation in agar autoclaving seems to be linked to autoclaving the agar with all other media components, especially the phosphates. Check this paper
Cathal, the ROS generation in agar autoclaving seems to be linked to autoclaving the agar with all other media components, especially the phosphates. Check this paper
Tanaka, T., Kawasaki, K., Daimon, S., Kitagawa, W., Yamamoto, K., Tamaki, H., Tanaka, M., Nakatsu, C. H. & Kamagata, Y. (2014). A hidden pitfall in the preparation of agar media undermines microorganism cultivability. Appl Environ Microbiol 80, 7659–7666.
Cathal (Phone)
Dec 14, 2015, 7:49:47 PM12/14/15
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Thanks for the paper Xabier!
Scott
Dec 14, 2015, 8:11:55 PM12/14/15
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Often worth while to stroll through expired or near expired patents to see how things were done. Here are a few relevant to this topic.
Algal strain for agar production
Stabilized agar product and method for its stabilization
Multi-differential agar culture medium
Production process of quick soluble agar
Agarose purification method using glycol
Low gel strength agar-agar
Cheers,
Scott
BraveScience
Dec 15, 2015, 2:38:39 AM12/15/15
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There's an article on g.xylinus cellulose matrix used as growth medium, and it showed much higher bacterial survival rate than agar agar.
And no inhibition.
And no inhibition.
Brian Degger
Dec 15, 2015, 4:59:32 AM12/15/15
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go go kombucha nerds :)
On Tue, Dec 15, 2015 at 7:38 AM, BraveScience <braves...@gmail.com> wrote:
There's an article on g.xylinus cellulose matrix used as growth medium, and it showed much higher bacterial survival rate than agar agar.
And no inhibition.
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Ilya Levantis
Dec 19, 2015, 11:25:47 AM12/19/15
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OK I'll bite. Can I have a link to growing cultures on BC though - do I simply grow the BC in a petri shape or blend before pouring into the petridish? My initial reservation is the porosity of the BC might make colonies less nicely defined. But hopefully I can give it a good crack and try a number of options.
Also RE:capillary electrophoresis.
I looked into this a couple of months ago and the literature trail for using agarose filled capillaries seemed to go dead after the late 80s - there seemed to be suggestions that getting the agarose to bind to the inside walls of the capillary was an issue. I suppose DNA moving in the gap between the gel and the capillary wall might prevent the formation of a clear band?
A 2004 review of different polymers used for DNA capillary electrophoresis:
Also RE:capillary electrophoresis.
I looked into this a couple of months ago and the literature trail for using agarose filled capillaries seemed to go dead after the late 80s - there seemed to be suggestions that getting the agarose to bind to the inside walls of the capillary was an issue. I suppose DNA moving in the gap between the gel and the capillary wall might prevent the formation of a clear band?
A 2004 review of different polymers used for DNA capillary electrophoresis:
Xu, F. & Baba, Y. Polymer solutions and entropic-based systems for double-stranded DNA capillary electrophoresis and microchip electrophoresis. ELECTROPHORESIS 25, 2332–2345 (2004).
DOI: 10.1002/elps.200405923
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