DIYbio 3 - Gel Electrophoresis
Mackenzie Cowell
Thanks to everyone for a great meeting: Michael, Jason, Sophia, Benny, Topher, Ricardo, Alex, Alec, nublabs, and everyone else.
Overview:
DIYbio 2 & 3 were focused on gel electrophoresis - we started by
researching all the amateur gel protocols we could find online (notably
the MacGuyver Project and the MAKE protocol, the latter of which we decided to focus on) and making a shopping list of materials
we would need to build the complete gel apparatus, make a gel, and
stain DNA. We got Agar agar powder from a health food store on ebay and
Potato Dextrose Agar and sybr green from a lab equipment reseller on
ebay.
Basically we over-stained the gels with methylene blue and couldn't visualize anything because the entire gel was a solid blue.
Also, we used metals that were way too reactive as electrodes in the
gel box and the redox reaction that they enjoyed probably ruined the
voltage field across the gel.
That said, I think we accomplished a lot, learned a lot, and somehow managed to do everything the protocol required in a spontaneous, parallel way - to me, that was the best part about the event. I remember stepping back at one point and being amazed at how well everyone was working together without any central plan: Topher and Jason were improvising with the stovetop and microwave to boil the agar, Sophia and Mike were cutting up charlie cards to make gel combs, and Benny, Ricardo, and the nublabs crew were constructing and taping gel trays and boxes. It was awesome.
What I learned:
1. We were not as rigorous with the materials as we needed to be. One
of our main problems was getting the concentration of the gel just
right, which was difficult because the packages for our agar didn't
list the original concentrations. In general, we followed the MAKE
protocol too specifically while using supplies that came with ambiguous
descriptions at best. DIYbio protocols we develop should be designed to
help the user understand the basic principles of the operation in a way
that enables them to improvise successfully with non-standard materials.
2. We should have had a solid understanding of how to dispose of all the materials involved before beginning as well as exactly what the potential risks involved were (very low - probably the biggest risk was spilling molten agar) and what to do in case something went wrong.
3. It would have been better to have built all the equipment in one meeting and then used it in a second. Trying to do both was a little too long.
Next week we are going on a tour of the Boston FabLab, courtesy of our friends at NubLabs - meet at the FabLab at 7:00pm on Thursday, 17 July 2008.
See you then!
Mac
p.s. go comment on the flickr photos!
p.s.s cross-posted to diybio blog: http://blog.diybio.org/2008/07/diybio-3-gel-electrophoresis.html
Mackenzie Cowell
Mac
Topher Cyll
Next time,
Toph
On Mon, Jul 14, 2008 at 11:14 PM, Mackenzie Cowell <maco...@gmail.com> wrote:
> Thanks to everyone for a great meeting: Michael, Jason, Sophia, Benny,
> Topher, Ricardo, Alex, Alec, nublabs, and everyone else.
>
> Overview:
> DIYbio 2 & 3 were focused on gel electrophoresis - we started by researching
> all the amateur gel protocols we could find online (notably the MacGuyver
> Project and the MAKE protocol, the latter of which we decided to focus on)
> and making a shopping list of materials we would need to build the complete
> gel apparatus, make a gel, and stain DNA. We got Agar agar powder from a
> health food store on ebay and Potato Dextrose Agar and sybr green from a lab
> equipment reseller on ebay.
>
> Basically we over-stained the gels with methylene blue and couldn't
> visualize anything because the entire gel was a solid blue.
>
>
> Also, we used metals that were way too reactive as electrodes in the gel box
> and the redox reaction that they enjoyed probably ruined the voltage field
> across the gel.
>
Bryan Bishop
> 1. We were not as rigorous with the materials as we needed to be. One
> of our main problems was getting the concentration of the gel just
> right, which was difficult because the packages for our agar didn't
> list the original concentrations. In general, we followed the MAKE
> protocol too specifically while using supplies that came with
> ambiguous descriptions at best. DIYbio protocols we develop should be
> designed to help the user understand the basic principles of the
> operation in a way that enables them to improvise successfully with
> non-standard materials.
One of the ways that the biotech toolkit has been solving this problem
is the start of a system that can help users acquire materials
[somehow] with very specific concentrations and other
characteristics -- the actual framework that could be used either by
suppliers or a way for users to properly tag products. And since not
everything is necessarily digital, the instructions for measurement and
analysis can be uploaded and implemented too. This sort of information
about what not to do (different protocol with different stuff) would
make for excellent use cases and quick writeups if given slightly more
detail within a specifically documented context. Remember, we're trying
to be sort of scientifically industrial here [while also having a fun
time :-)], but being less than exact here and now can produce pains in
the long run in this community. What I mean by all of this is to start
off with ridiculously simple XML files. Let's go code some class
structures and implement objects to represent protocols. Include some
variables for human description of the steps, and have that print out
information when we decide what it is that we want to do from our
knowledge base. In the future it would be easy to subvert parts of this
computer code with physical automation hardware (for instance, USB plug
into an Arduino into the thermocycler -- you don't have to run through
the crappy menus any more, hurray, and less chance for human error,
hurray).
> 2. We should have had a solid understanding of how to dispose of all
> the materials involved before beginning as well as exactly what the
> potential risks involved were (very low - probably the biggest risk
> was spilling molten agar) and what to do in case something went
> wrong.
I think some of this information could be integrated with the MSDSes. As
for the actual training, I'm not sure how far you want to go with that,
especially if you refer to large group work. It sounds like you had 10
or 15 people show up and help out? I'm not even sure how many labs
coordinate that many people in realtime. ;-) It requires some amount of
rehearsal, no doubt.
> 3. It would have been better to have built all the equipment in one
> meeting and then used it in a second. Trying to do both was a little
> too long.
Since this all has to be repeatable, do we have the exact specifications
of the equipment? Any details on the logs being kept? Anything like
this whatsoever? Or are we all just dust in the wind?
Either way, sounds like it was fun. I'm glad.
- Bryan
Uhh. My server is down for an unspecified time. Maybe someone would
upload the toolkit to diybio.org until I get back up and running? Might
end up just being a few days. The server was getting somewhere near
300,000 hits a month, and we're losing valuable eyes here ...
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