Re: [DIYbio] awesome beginner level biotechnology projects anyone?

[Jeswin]

Welcome to the community. I know what you mean.

Anyway, I think we [diybio community] should have some sort of
distributed lab-work setup. If people like Annie can volunteer lab
time, then maybe small projects can be completed.

On Sun, Jan 6, 2013 at 4:34 AM, R Annie O'NieL
<aliasano...@yahoo.com> wrote:
> hello..the first thing that popped into my mind when i saw this site is
> utter amazement..im so glad i found this site.You should be very proud of
> your site. I am currently doing my bsc in biotechnology in India,Goa.
> Although i had an extremely bad depression phase when i didnt get a seat in
> medicine because of petty government policies and the eternally damned
> system...this does help my coping mechanism...to interact with fellow bio
> enthusiasts...as not many in my class are as interested in biology as i
> am...I would love to share as much as information as i can, and gain some in
> the process..i have access to my lab at school, and am planing on using it
> for simple projects... I am also interested in methods to preserve
> environmental health...synthesising organisms that can degrade different
> varieties of plastics, various forms of biofuels...etc...I look up to your
> expertise, knowledge, guidance and experience.. please do suggest some
> interesting beginner level projects i can try out at college when i am
> free..beats sitting in a bench alone, bored to tears...p.s sorry for the
> grammatical errors..
>
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[Sebastian S. Cocioba]

If anyone in the NYC area is in need of a plant tissue culture lab I can offer my space, equipment, and expertise at cost of reagents used. If the project is small or one cannot afford to pay Ill just waive the fee. Im set up in woodside and have tons of PTC experience. The place is small but functional and close to mass transit. All are welcome! :)

Sebastian S Cocioba
CEO & Founder
New York Botanics, LLC

Sent via Mobile E-Mail

[Mega]

Sebastian, that sounds awesome!!

I'd definitely visit it if I lived in NYC... ;) 

[Mega]

R Annie O'NieL  (Annie is Ok?) , Hi!

Great to hear that!!! 

Forget medicine :D 

Imo, medicine will, sooner or later, be replaced by genetic therapy which will make costly-producing medicine pointless. 

.as not many in my class are as interested in biology as i am..

You surely are not the only one here ;) 

For many students it's just a means to get a job,   when they leave university in the afternoon/evening  they no longer think about biology.  

Which semester are you? 

Oh, I'd reccomend you an experiment, if you like bioluminescence: There is a plasmid called pVIB which, when carried by E.Coli, makes it glow in the dark. I bought the plasmid privately (they sent it to my home adress without prroblems). The plasmid itself costed 10 US-$ (I then ordereed two pieces, in case one would be spilt/lost)  , the shipment cost was some 30$ though. 

Best, Andreas

[Mega]

Here're pictures how it should look like ;)

For the bioplastic thing, I remember George church talking about that. He's a co-founder of many biotech companies, including biofuel from rotten wood, from sunlight; etc.

[Mega]

http://diyspartanbiotech.wordpress.com/2012/05/11/transforming-e-coli-with-pvib-plasmid/

Here's the link, I forgot before :D

[Andreas Sturm]

You're welcome ;)

He's probably one of the most influencing biologists that are still alive :)

On Sun, Jan 13, 2013 at 7:21 AM, R Annie O'NieL <aliasano...@yahoo.com> wrote:

ome...will definetely look it up...forgive me for my ignorance,but i never heard of george church:(....will definetely look it up tho,its bound to b sumthin fabulous:)...thnx for the pics they look amazing...:)

[Andreas Sturm]

*probably one of the top 10 or even top 5

[Joro]

yes please .
Sent from my ASUS Pad

Andreas Sturm <masters...@gmail.com> wrote:

*probably one of the top 10 or even top 5

[Mega]

Haha :D Yeah, that'd be nice ;) 

I've got his book sometimes ago. 

 met him in the Ars Electronica, and it was very interesting. 

[Nathan McCorkle]

On Sat, Jan 19, 2013 at 4:18 AM, Kwabena Owusu-Boateng
<ghcop...@gmail.com> wrote:
>
> Hi! I must be living under a rock or something, cause all my teachers tell
> us here in Ghana is that your biotech degree enables you to work in lab
> analyzing blood samples or the the beverage industry.

Sure, you /would/ be able to work in those industries, but it could
also prepare you for much much more.

>Happy to be involved
> in this community. Interesting ideas too!

Please stick around and post often!

--
-Nathan

[Andreas Sturm]

Welcome ;)


Well, it's just... you know, teachers are also just humans. And if they have never heard of diy bio, they can't tell you anything about it...

[Kwabena Owusu-Boateng]

Hello again,

Can anyone help me with a simple plant transformation experiment with plants, equipment and techniques will be helpful.

[Koeng]

Me too! That would be awesome. I want to get into plant transformation

[Andreas Sturm]

Hi!! 

Do you have access to agrobacterium? 

Or a university that can get you some? 

If not so, you can use this technique:

http://www.jove.com/video/2560/efficient-polyethylene-glycol-peg-mediated-transformation-moss

PEG you can get from the internet as "Miralax" or omething like that. 

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[Sebastian S. Cocioba]

Miralax is PEG 3350 and the generic forms are just as good. The publications call for a range of PEG molecular weights from 1300-8000 with varying results. It seems to be dependent on the net quantity of each lipid type in the plasma membrane of the plant cell which also varies between species. I would say to start with cheap miralax and then tailor the protocol. You will however need enzymes which tend to be costly. Im going to post a new thread right after this regarding Cathal's suggestion of store bought enzyme capsules and their efficiency ad protoplasting agents. 

Sebastian S Cocioba

CEO & Founder

New York Botanics, LLC

Sent via Mobile E-Mail 

[Kwabena]

Is there are way you can transform a plant to change colour in say, soil with a high concentration of heavy metals?

[Dakota Hamill]

If you look in the parts registry under promoters

http://partsregistry.org/Catalog

You can find heavy metal induced promoters.  Throw another gene downstream of that, like GFP or maybe a lux operon, and you could have bacteria or yeast that would glow in the presence of the heavy metal.  Spray it over a field or contamination site and check for the presence of your expressed protein.  Easier said than done though.

You could also make it work in plants like you say, though it would probably take a lot of work.

One thing that would be very important is the concentration at which the promoter is induced.  If it is SUPER sensitive, than your biological alarm system might be making you think your soil is more contaminated than it is....or it's still at safe levels, but enough to turn on the genes.   

I think living bio-sensors are probably one of the coolest things one could design with the biological parts, and they could have a lot of commercialization potential I'd think.

Too bad none of us can get parts though.

[Sebastian S. Cocioba]

You could take that promoter and tie it to an anthocyanin pigment protien gene and any white flower will begin to change color to red...theoretically. The antho genes are different for each species but not by much. The plasmid construct may be a bit hard but the transform is easy. Now just find a white flowering plant thats not weedy or invasive and plant. There are all kinds of regulations to pass for it to be approved. In the US its an EPA field test that takes two years and them some paperwork. It can be done.

Sebastian S Cocioba

CEO & Founder

New York Botanics, LLC

Sent via Mobile E-Mail 

[Kwabena]

Wow! This is interesting stuff but it could take months to complete even if i can get access the proper equipment  I was just looking for a project that can be completed within a month and can rock a science fair. Sebastian when you say 'theoretically' does that means it cant work in the real world? Dakota, you seem to know a lot about this, have you done any work in that field? 

[Sebastian S. Cocioba]

When I said theoretical i mean't that to the best of my knowledge no one has tried this yet. The experiment, in parts however, has been done. There are studies on the exact genes of pigmentation in flowers where anthocyanin is highly conserved among many species. Others have studied abiotic stress with relation to heavy metals and IIRC have Identified some heavy metal sequestering pathways in algae. The eukaryotic version of bacterial heavy metal promoters might be an issue if you cant get access to a particle gun. Chloroplasts, as you may know, have prokaryotic expression machinery so there would be no need to mod the bacterial promoter. Agrobacterium only transfers into the nucleus. PEG may work (I'm developing a protocol for that atm). Either way its a long term project.

If you want to just kick some bootay at a sci fair, why not stick to bacterial stuff since the turn-around time is way shorter than anything plant related. Is the fair local, regional, national? How would you describe the expected complexity of the experiments that someone would enter into said fair? I would not go overboard if its just a school thing...unless of course you want to. Once you start doing biotech stuff, its hard to stop.

Sebastian S Cocioba

CEO & Founder

New York Botanics, LLC

Sent via Mobile E-Mail 

To view this discussion on the web visit https://groups.google.com/d/msg/diybio/-/7T-CPFI9s7kJ.

[Dakota Hamill]

> Dakota, you seem to know a lot about this, have you done any work in that field?

Short answer, no. There are a lot of people on here with way more
experience doing hands on molecular biology than me. I discovered
iGEM a year or two to late to try to establish a group at my school,
and coming from a small state school with non-existent research
departments and faculty who do no research, gaining hands on
experience in advanced molecular bio wasn't as easy, and so became an
after school thing. Lack of funds and equipment, although sometimes
depressing, should never deter someone from pursuing what they want to
study, or achieving a dream or goal. Whenever I feel like research is
impossible because I lack this or lack that, I try to think of all the
great scientists in the past who have done amazing things with so very
little, and it all seems possible again.

I don't know what situation you are in in terms of access to lab
equipment, but generating ideas is always free! Spending time in the
parts registry was fun because you'd start to see a coding protein
part from one project, then a really cool promoter from another
project, and think...hmm combining these would make a great new
biobrick!

Brainstorming new biological parts is really neat once you have a
basic understanding of what is needed in a plasmid, because the
plasmid becomes your blue print, and you start to view the E. Coli as
a billion little factories that build their own machinery, repair
broken machines themselves, and the only thing you need to pay them
to make your blueprint is some sugar and basic nutrients! (Forgot to
mention, they also build brand new factories by copying themselves
every 30 minutes!)

But ideas are easy to come by, execution of those ideas into a viable
tangible thing is what takes time, effort, and money. Take advantage
of being at University, look into maybe starting an iGEM team if you
can find a passionate professor who will oversee the group.

I've since moved away from bio-brick stuff simply because it's a pain
in the ass to get access to anything, and too expensive to synthesize
yourself. Maybe if I return to school I'd pick it up again, but for
now I'm focusing on easier molecular bio stuff and natural product
isolation.

Being in Ghana, you have a lot of access to a ton of interesting plant
life with amazing secondary metabolites! If you're interested in
chemistry I could help you in that department.

If I had a month to do something and I was in Ghana, it'd be this;

1. Identify plants that live in hostile or unusual circumstances, or
plants that have a history of use by the local culture as sources of
medicinal qualities (study known as ethnobotany)
2. Gather leafy tissue (or roots / bark in some cases) and grind the
tissue in a mortar and pestle.
3. Get a seperatory funnel and extract the tissues using a solvent
such as ethyl acetate, methanol, or dichlormethane.
4. Collect your fractions, one fraction is non-polar (ethyl acetate /
DCM) and the other fraction is polar (methanol / water)
5. Now whatever natural products and secondary metabolites of
potential medicinal quality are useful in your plant, are now either
in a polar or non-polar solvent depending on their molecular
structure.
6. Use a rotary evaporator to remove excess solvent so you are left
with a crude plant extract. If you do not have a rotary evaporator,
heating gently on a hot plate under a fume hood will remove solvent,
but much more slowly than a roto-vap. Be careful not to use to much
heat as you may decompose some of your products at to high a temp.
7. Re-suspend your crude plant extract in the least amount of solvent
possible (to keep it concentrated) 1-3mL
8. Put some of the plant extract on filter paper discs, and perform a
disc diffusion assay against a few different organisms.

http://en.wikipedia.org/wiki/Kirby-Bauer_antibiotic_testing

Gram negative - E.Coli
Gram positive - B. subtillus
Fungi - Aspergilus / Candida / Sarchomyces

Timeline:

Selection of plants based on literature of past use, or local culture
use = 1-5 days

Collection of specimens in the wild = 1-5 days

Grinding of tissue and solvent extraction per plant = 1-3 hours per
plant sample, so multiple per day. Add 1-2 hours if you don't have a
rotary evaporator. If you are using dichloromethane it could
evaporate out over night at a warmish temperature in a flow hood. The
higher the boiling point of your solvent (ethyl acetate or water) the
longer it will take to remove it. Methanol / dichlormethane can be
removed in short time. You can't "boil" out the water because then
there is a high chance you are completely destroying whatever
anti-fungal or anti-bacterial properties the compound has. (think
penicllin)

Plating of bacterial samples = 1 hour

Growth of bacterial samples + disc diffusion test = 1-2 days
(overnight) for bacteria, week+ for fungal

Theoretically, you'll hopefully have some rings of inhibition of your
filter paper discs, showing you that compounds in your plant extract
are killing or haulting the growth of certain types of bacteria or
fungi.

That's the easy part!!! Now separate the crude extract with HPLC,
fractionate it, prep on a normal phase column, do LCMS, GCMS, and NMR
of the fractions, and discover a brand new drug!

But seriously, Africa is full of amazing plants with great potential,
and you have a good opportunity to get real science data, and the best
part about it is that you don't need crazy expensive equipment to do
these extractions + disc diffusion assays. Think of it, shamans and
healers over the millenia have made extractions of natural plants,
only they don't do a disc diffusion assay to kill the bacteria, they
just put it on the open wound or cut!


A quick google search shows 3 people at your school, KNU (sorry...to
hard to keep looking up the spelling haha) wrote a book on medicinal
plants. I generally have access to ACS stuff, but not book chapters I
guess.

T. C. Fleischer, , M. L. K. Mensah and , R. A. Dickson

http://pubs.acs.org/doi/abs/10.1021/bk-2009-1021.ch014


Maybe go talk to this dude? He's at your school and maybe he can help you

http://www.knust.edu.gh/pages/sections.php?siteid=pharmacy&mid=600&sid=2867&id=658

If you have any questions I'll answer them to the best of my abilities.

[Kwabena Owusu-Boateng]

Thanks a lot Dakota big help.