Re: Injection of GFP into Kyoto Moss Spores: Suggestions?

[Mega]

Hi,

Well, moss is a eukaryote and "we" Eukaryotes usually don't carry plasmids (except of yeast, no e. plasmids are known AFAIK)

So you'll need to insert the gene  a)  into the plant's nuclear genome  or   b)  into the chloroplasts.

a) is much easier, doesn't need homologous recombination.

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

Here they did exactly that.

Or you use agrobacterium. That is far more easy, but it is difficult to get the Ti-plasmids needed for non-academic labs.




On Thursday, November 29, 2012 3:17:43 PM UTC+1, Jeremiah Blondin wrote:

Hello Everybody.  Nice to meet you.

I recently became involved in DIY biology. I am currently working with a thermal cycler from OpenPCR, dremelfuge, a 4 cell water bath, a cheap plastic electrophoresis tank from Teachers' Discovery, and an old but still operational incubator that I picked up at a lab auction. I  have been wanting to move beyond E. Coli experiments for some time.  Recently I decided to try injecting Green Fluorescent Protein into moss spores for use as decoration in the cage of my frilled dragon, Helix.  I got the idea after watching a lecture by the DIYbio organization in London where moss was suggested as a possible alternative model organism.

So, Here's my question. I have done GFP injections in E. Coli.  What modifications would be necessary, if any, to perform the procedure on moss?  I have ordered Kyoto Moss Spores from Amazon so, assuming that their viability, this is the specific variety that I will be using.  Any suggestions?

[NB]

Not to discount the previous post, which looks to be a very reasonable technique (one I'd like to try), but 'we' eukaryotes are not ostracized from the plasmid scene. We even have linear ones.

Plasmids of Eukaryotes - Fundamentals and Applications

Book review

http://onlinelibrary.wiley.com/doi/10.1002/jobm.3620270912/abstract

Google Books

http://books.google.com/books/about/Plasmids_of_eukaryotes.html?id=fLMTAQAAIAAJ

Linear plasmids among eukaryotes: fundamentals and application

PDF

http://link.springer.com/article/10.1007%2FBF00312851?LI=true

Cheers,

Nick