No need to apologize. I think most of us have no idea on how to get
started since not all here are in the bio/chem field.
> interested in doing some home experimentation in biotech. I'm a final
> year applied maths PhD student and would like to think I know the
> basics, the theoretical basics that is, of biotechnology in the modern
> age. My interest is in RNAi and transient gene expression through
> insertion with Adeno-associated and Adenoviruses and Lipoplexes. This
> is in the context of morphogenesis and homeostasis in polycellular
Seems like you know more than you think. Think of yourself as a brand
new grad student and how would you go from there. The people around
here can advise you on where you should go. Judging from your area of
interest, which I don't know much about, this is not some trivial
subject like "how to grow GFP __{insert plant name}____". Without
doing some research, I guess you are dealing with gene insertion with
viruses. Seems like something done in a university lab. Can this be
done at home? We're talking about viruses here so...
> eukaryotes. Most probably samples taken from my self.
Whoa, working with mammalian cells is on another scale in the first
place. Working with your own cells might be disastrous. What if you
culture your cells and you change their DNA. Then you accidently get
those cells into your own body. Bad things are going to happen. I
haven't worked with mammalian cells before and safety-wise, I think
they're on a different level than bacteria or fruit flies.
> What I lack is any kind of experience with the experimental side of
> biotechnology. What sort of lab can I expect to build oriented to that
> sort of stuff. Funding wise my budget is in that grey zone of being
> more than the average hobbyist can afford but a good deal less than a
> well funded university lab. Can any one advise me on what I might need
> to get started here?
Labs are not cheap. Even with used equipment, you're probably looking
at thousands of dollars. A nice microscope is at least $100. Also,
depending on the subject you choose, the price will go up.
You're applied math. Why not start with some modeling? Can that be
done? It would be cheaper and less troublesome. I am very interested
in modeling but have very little experience with that.
Good luck. You might want to reconsider the self-samples part though.
-JJ
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Whoa, working with mammalian cells is on another scale in the first
place. Working with your own cells might be disastrous. What if you
culture your cells and you change their DNA. Then you accidently get
those cells into your own body. Bad things are going to happen.
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--Incubator with non-dry environment... you could build this yourself,
but unless you know how to build electronics and program, you can
probably find one at an auction or on ebay. (as for the rest of the
equipment)
--laminar flow hood - again you can build this, there are guides
online... some as jerry-rigged as a clear plastic tub and a HEPA
filter siliconed to a hole cut in the backside. UV light inside it
turned on when you aren't doing work is good measure IMO too.
--pipettors, pipets syringes and fine needles (a lot of antibiotics
come in sure-seal bottles), scalpels (you mentioned tissue), culture
dishes and multi-well plates, inverted stereo microscope
--growth media (MEM, DMEM, RPMI, serum-free), serum (bovine, fetal
calf, horse, goat, probably your own??), antibiotics/antimycotics
(streptomycin, penicillin, amphotericin)
(google 'DMEM antibiotics', first 4 links deserve a reading)
--cells - can be ordered as live mice with known genotype, frozen cell
lines (often cancer cells are of use), extracted from live organisms
I don't think eating chicken or eggs is illegal in the UK, so if you
bought a live chicken and cooked dinner (as many people do) with all
but the heart, which you threw in some DMEM to keep going, people may
think you're weird but there should be no fuss IMO... you could pierce
your ear, and throw the chunk you remove into some media likewise, how
else were you thinking of getting your own cells?
--
Nathan McCorkle
Rochester Institute of Technology
College of Science, Biotechnology/Bioinformatics
That is so true. Unless you did something in a lab, you won't truly
appreciate how hard it is to get something as simple as bacterial cell
cultures to work. Sometimes the skills are not really lab related like
steady hands and keen eyesight. You know how hard it is at first to
fill electrophoresis gels wells without contaminating nearby wells?
You got a PhD in applied math but that is as practical when it comes
to lab work as a PhD in EE is.
Maybe you can get a teaching post for a lab class? I know the TAs
usually try to reproduce the experiments beforehand or set it up. I
don't know about your University but you could try.
Ambition is admirable, and I'd encourage you to follow your ambitions to the fullest extent, provided you're aware of how much work and risk is involved. You may not be able to jump into advanced cell culture without working first with simpler projects to learn key skills, but if you're determined, it's your learning curve. :)
I'd also encourage you consider and revise your plans according to hazard and legality: not everything is appropriate to DIYbio in the same way that not everything is appropriate for hobby electronics tinkering (unlicensed radio, high voltage, and driving homemade vehicles on the motorway without a license come to mind).
There's a point, in other words, when your project is either hazardous or illegal enough to warrant finding alternate means to do it. You could still do it yourself (DIY), but it may hotel getting a license and building a licensable premises. I've hit this wall in Ireland, where you must get a license to work with GMOs of any sort, no matter how inane or innocuous the alterations. So, I'm getting one. If it's possible you need a license where you live to work with potentially infectious human cell lines, make sure you've done all the research.
Regarding self-biopsies.. I'd suggest having them cut out entirely rather than shaving or punching only a little. Pre malignant growths can be stimulated to full malignancy by biopsy, so for small things like moles it's better to chop than shave.
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> You sound pretty keen for a math maj...
> On Fri, Apr 8, 2011 at 9:04 PM, CodeWarrior <code.w4rr...@gmail.com> wrote:
> > Another thought. I...
> > For more options, visit this group athttp://groups.google.com/group/diybio?hl=en.
>
> --
> Nathan McCorkle
> Rochester Institute of Technology
> College of Science, Biotechnology/Bio...
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There is a lot of filtering that you would have to do, but is blood
enough to grow cells on? Could you "feed" your own experiments? If
that is too gruesome, what about getting it from a flock of healthy
goats? It seems that the only bad thing about this is that using a
needle everyday isn't healthy for the animal's (or your) blood vessels
and thus decrease the overall fitness of your blood source (which is
not cool, because goats are friendly, and you don't want to put
yourself at undue risk).
It seems like DIYbio should think about easy ways to produce feedstock
for these more touchy cells... even for minimal media recipes for
bacteria, there are a lot of special components. Maybe some sort of
transgenic yeasts or bacteria that secrete only certain amino acids,
so a cell free growth media would have /mostly/ one amino acid.
Purifying the salts seems like the only next step to having pretty
pure aminos... mix them with some phosphate buffers and serum (which
would be acquired some other way).
Our bodies make everything our cells need, so a bio-fermentor solution
to this problem IS possible.
--
Insect cell culture interests me, but I know nothing yet of it. Based solely on the fact that few intracellular parasites (viruses, bacteria) exist that shuttle between human and Insect hosts, Insect cell culture makes a lot more sense.
I do think that, by the time you have established a DIYbio cell culture lab that works, you could probably have completed a college degree in cell biology and have gotten a position studying it somewhere better equipped, possibly for the same or less cost. Some things aren't well suited to DIYbio just because the economies don't make sense, although there's a case to be made for do-it-with-others bio when this is the only barrier.
It stands out at me though that plant cell and tissue culture is an untapped area for us: it's not much harder than bacterial culture and it can be more practical and more rewarding than mammal or insect because you can optionally regenerate entire plants from your experiments. Indeed, it's often used to mass propagate valuable isolates industrially.
I studied plant tissue culture and biotech in college and have a little practical experience, so I'd be able to offer some pointers to people with an interest in this area, although there might be someone else lurking around here with true practical experience!
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On 13 Apr 2011 07:43, "jlund256" <jlun...@gmail.com> wrote:
> There is a lot of filtering that you would have to do, but is blood
> enough to grow cells on? Cou...
No, no human blood, a hundred times no. Same with livestock. Blood
serum is a common additive to mammalian cell culture and is collected
from slaughtered animals. Usually as an additive at 5% or 10% or
media volume to provide protein growth factors and whatnot. That
would be the source, but really, mammalian cell culture isn't worth
the trouble.
Insect cell culture probably isn't worth the trouble either, but is
cheaper/easier/safer, the provides most of the cell types, genes, and
biology happening in mammals.
Or try a squid. I hear they have neurons big enough to stick a straw
into.
> It seems like DIYbio should think about easy ways to produce feedstock
> for these more touchy c...
There's a strange interest on this list in taking existing lab
protocols, and replicating them as DIY projects by replacing the
reagents known to work with baroque and complicated DIY replacements.
Well, these are mostly thought experiments, or just notions. These
proposals nearly always seem unlikely to work and difficult to
implement. You would spend months and years assembling ingredients
and never bake a cake.
Instead, I suggest looking for interesting things to study and/or cool
experiments to do, and find the simplest and fastest way to do them.
The best plants and animals to study are those at close at hand.
Jim Lund
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It seems like DIYbio should think about easy ways to produce feedstock
for these more touchy cells... even for minimal media recipes for
bacteria, there are a lot of special components. Maybe some sort of
transgenic yeasts or bacteria that secrete only certain amino acids,
so a cell free growth media would have /mostly/ one amino acid.
Purifying the salts seems like the only next step to having pretty
pure aminos... mix them with some phosphate buffers and serum (which
would be acquired some other way).
Our bodies make everything our cells need, so a bio-fermentor solution
to this problem IS possible.
I studied plant tissue culture and biotech in college and have a little practical experience, so I'd be able to offer some pointers to people with an interest in this area, although there might be someone else lurking around here with true practical experience!
At a cellular level immortalisation in insects is probably pretty similar to mammals. Mostly it's a matter of providing telomerase, I think?
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On 13 Apr 2011 17:45, "General Oya" <gener...@gmail.com> wrote:
We're currently culturing SF-9 insect cells (on passage 10 in T-225's). If we were licensing them as compared to using them in an educational environment, we'd have to pay Life Sciences around $75,000 a year just for the priviledge.
I'd love to culture some Golden Orb Spider spinnerette cells... but I'll still need to figure out how to immortalize, unless the Chinese have already created a Master Cell Line.
I think we'll need to figure out a few things about mammalian cell culture if we really want to produce functional bio-printer projects. Once again, I've found the Cell Culture for Tissue Engineering text to be quite helpful.
JCVI may actually be on the precipice of building a community lab capable of Cell Culture here in Maryland. We'll have to see how this project develops.
Ryan
On Wed, Apr 13, 2011 at 6:05 AM, Jonathan Street <streetj...@gmail.com> wrote:
>>
>> It seems...
I guess what I meant is that the body takes easily acquirable
feedstock and turns it into everything our bodies need. So imagine if
you could take an animal with the smallest genome, and start
attempting to strip off working genetic modules... there'd be no need
for all the macro sensors like eyes, ears, touch, smell, but the
immune system and skin and ECM would be good things to keep, and maybe
a mouth too, with a mastication inducer as well.
And for the non-essentials, some other organism produces them, so pull
in those operons/modules and forget about it being a problem.
Alternatively, if sequencing and synthesis were in the DIY realm, we
could just do all this in vitro. Bootstrap with some purified ribosome
and RNA polymerase, and you could theoretically have an in-vitro
"cell" or "organism"... conveniently with digital control systems.
--
sure if that's what you want to do... if you want to work for
yourself, and be close to your community, what I am speaking of would
be something like a sustainable lab co-op... you wouldn't have to deal
with Life Sciences $75k/year licensing type things... I think this
could only be possible by using bio-tech to our advantage, by having
it do more work and allow us to rely less on preparative chemistry
techniques for purification.
doesn't blood infuse all the nutrients into the tissue it passes?
couldn't you then just dialyse blood to leave all the big stuff
inside, and enrich the dialyte with serum? (this is a real question,
not rhetorical)
> Insect cell culture probably isn't worth the trouble either, but is
> cheaper/easier/safer, the provides most of the cell types, genes, and
> biology happening in mammals.
>
> Or try a squid. I hear they have neurons big enough to stick a straw
> into.
>
>
>> It seems like DIYbio should think about easy ways to produce feedstock
>> for these more touchy cells... even for minimal media recipes for
>> bacteria, there are a lot of special components. Maybe some sort of
>> transgenic yeasts or bacteria that secrete only certain amino acids,
>
> There's a strange interest on this list in taking existing lab
> protocols, and replicating them as DIY projects by replacing the
> reagents known to work with baroque and complicated DIY replacements.
> Well, these are mostly thought experiments, or just notions.
Brute force is how knowledge has been acquired since the beginning of
time. Linux isn't always as plug-n-play as Windows, but those
programmers who built it just didn't want to pay for licensed
technology they couldn't take apart or have the potential to improve.
> These
> proposals nearly always seem unlikely to work and difficult to
> implement. You would spend months and years assembling ingredients
> and never bake a cake.
sure, but there is something to say for that, because what if after
that time you are really good at baking lots of cakes? Would you buy
cakes from a baker that used all local ingredients and had wholesome
business practices?
Even if you did this without some crazy scheme using crazy unheard of
systems, you don't think its possible to beat those prices?
>
> Instead, I suggest looking for interesting things to study and/or cool
> experiments to do, and find the simplest and fastest way to do them.
> The best plants and animals to study are those at close at hand.
>
> Jim Lund
>