I use agro strain GV3101 pmk90. You could probably have it shipped to you if you write to Stan Gelvin's lab over at perdue. My go to transformation plasmid is pPZP-200 series that confers kanamycin resistance as a selection marker. Floral dip sucks in my oppinion and youll be limited to arabidopsis only. Most important part of plant genetic engineering is tissue culture. You can build a laminar hood for cheap using a hepa air purifier, giant clear plastic storage bin, and some ducting. That way you could readily transform any plant (YMMV).
Just put a plant promoter like corn ubiquitin or CaMV in the mcs, them your egfp, then a terminator like the nopaline synthase one already in your plasmid downstream in the selection marker region. Then transform agro with the plasmid. Strain gv3101 is aggressive but a little hard to get rid of. It has genomic gentamycin resistance and spectinomycin in the plasmid if you are using pzp. I can send u my plasmid if you cant get it elsewhere. I transform tobacco on a daily basis and its my goto model organism. I can also send you seeds if you want to join the cult of nicotiana. Ive made some fluoroplasmids to shoot with my gene gun and results were fair. I use non optimized cometgfp and could never get my hands on egfp. Ill trade you my agro vector for yours with egfp. 😃 also feel free to ask whatever you want. I like helping plant newbies. I post all my lab stuff on fb and twitter. More than welcome to follow if you'd like. Good luck!
-Sebastian
Sent from my iPad
-Sebastian
Sent from my iPad
I'm currently an undergrad studying molecular biology, and I recently joined a community lab after taking biochemistry lab at my school. In my class, we transformed a colony of E. coli using egfp and I thought it was really cool, and I've kind of been obsessed with trying it with other organisms as well, so I wanted to see if maybe I could do the same thing with a plant. After spending a few days at the library and searching through a couple of scientific journals, I found that I could do a similar procedure with agrobacteria, which can use a disarmed tumor-inducing plasmid to inject a segment of tDNA from another binary plasmid into a plant genome (I'm still not 100% sure how that works or where in the genome it gets incorporated, but I know it works at least). I would like to try and put egfp into a plant and have it expressed at detectable levels. I've had people tell me it's not too difficult to do. If any of you have done it before, or if you know what is involved, could you possibly give me some pointers on how/where I can order the appropriate plasmids, and which strain of agrobacteria I can use? I found a couple of protocols, so I have a general idea of what I'm gonna do (floral dip), but I'm still a bit iffy on how to select the right plasmids and bacteria, and where exactly on the binary plasmid I should ligate my gene of interest. Any help is much appreciated :).--
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