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Matt Tucker

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Jul 27, 2017, 3:38:34 PM7/27/17
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After endless research and reading about genetics, I have finally decided to yield to my curiosity, and put some time into the lab. As such, I'm trying to find an affordable source for GFP to be used in a project where, agrobactarium- t. is used to deliver GFP into a plant. Thoughts?

Skyler Gordon

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Jul 27, 2017, 5:11:56 PM7/27/17
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Have you already obtained Agro. spp.? I would look into that before you start trying to get all the other materials.

-SG
On Thu, Jul 27, 2017 at 12:38 PM Matt Tucker <mathew....@gmail.com> wrote:
After endless research and reading about genetics, I have finally decided to yield to my curiosity, and put some time into the lab. As such, I'm trying to find an affordable source for GFP to be used in a project where, agrobactarium- t. is used to deliver GFP into a plant. Thoughts?

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Matt Tucker

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Jul 27, 2017, 9:01:50 PM7/27/17
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Skyler Gordon

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Jul 27, 2017, 9:17:49 PM7/27/17
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If you have the capability to buy plasmids, you can most likely buy one that already has GFP cut into it. Other than that I would ask some educational facilities if you can get a sample of the GFP gene to express yourself. If you can't find a plasmid that has GFP and the expression system you want, you may be able to buy the gene from ThermoFisher.

Many companies that sell plasmids won't let you purchase anything under a 'John-Q Public' account so you'll need a business account, non-profit would be the best way to go.

-SG

Skyler Gordon

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Jul 27, 2017, 9:21:42 PM7/27/17
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The gene itself is only about 240 bp, so if you can't find it for sale you might be able to build it yourself from long primers.

Mega [Andreas Stuermer]

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Aug 1, 2017, 10:20:43 AM8/1/17
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pCambia1302 already contains GFP, ready to use

Matt Tucker

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Aug 13, 2017, 3:12:28 PM8/13/17
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How about luciferase?

Skyler Gordon

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Aug 15, 2017, 8:49:57 PM8/15/17
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Luciferase is going to need other proteins to function and is also going to consume ATP to cause the glow it creates. GFP is a better route because it has fluorescence and doesn't just glow/produce light like luciferase would

-SG
On Sun, Aug 13, 2017 at 12:12 PM Matt Tucker <mathew....@gmail.com> wrote:
How about luciferase?


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a a

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Sep 9, 2017, 11:11:45 PM9/9/17
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Correct me if I'm wrong, but wouldn't luciferase be preferable if it didn't rely on flourescence to glow (doesn't need uv light)?

Skyler Gordon

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Sep 10, 2017, 1:13:35 PM9/10/17
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To be completely honest in this particular setting neither of these proteins will show any presence in the plant unless you're a whiz with protein signaling.

If you can get luciferin to be expressed in the right tissue, correctly fold, and then maybe be stored somewhere (some vacuole or empty lysozyme) you'll still run into the issue of having to feed it ATP to cause the glowing. And even then, it will glow constantly.

(Check out pyrosequencing if you want to get a detailed description of how luciferase works)

There is the possibility that you can control the expression via a gene signaling system like the Lac operon, but I'm not overly familiar with plant systems.

You could add the gene several times to make sure it's in the correct section of the genome (Agrobacterium are somewhat undirected in their insertion methods) and try to include a genetic signal that causes expression / is turned on only in the flowering tissue.

GFP is most commonly used as a protein conjugate in order to observe where the protein is located in the cell. So the choice is really going to depend on what you're using it for.

When you are trying to just make something glow for the sake of curiosity / wonder I would suggest trying to use as little ATP as possible. For the health of the plant.

I hope this helps.

-SG

Skyler Gordon

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Sep 10, 2017, 1:14:55 PM9/10/17
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* correction: luciferase will glow constantly as long as it has an ATP source. If fed intermittently it will flash

a a

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Sep 11, 2017, 8:50:00 AM9/11/17
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Autoluminescent luciferase based plants already have a proof-of-concept, and there are commercial plant-expressing luciferase plasmids. Here's the article: http://journals.plos.org/plosone/article?id=10.1371/journal.pone.0015461
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