The production of a trait may be many different genes (not sure if multiple produce thioalchol). Also, Staphylococcus hominis is a prokaryotic cell, and therefore the procedure would be much different than if you wanted eukaroyote-eukaryote. There is going to be a much different procedure each time, but most of the time you can check out KEGG for pathways. Perhaps, for example, take a look at http://www.genome.jp/kegg-bin/show_organism?org=sab to get BLAST comparisons. Also, I'm pretty sure that they've figured out the traits that produce thioalcohol ( https://www.sciencedaily.com/releases/2015/03/150330213947.htm , wikipedia link). You want to replace thioalcohol with ester, so naturally you want insertion to be over the thioalcohol. Alternatively, you can just replicate as a plasmid, albeit with less stability. Would allow you to get rid of the 'homology' parts of design below.
-- Your description of replacing the gene that produces thioalcohol with one that produces ester is not a project of identifying and isolating genes but unfortunately one of cloning, transforming, and troubleshooting. Not quite as exciting. No identification is needed (since they've found the genes) and isolation is a vague description (deleting, transcriptionally isolating, isolating from genome, orthogonal isolation, or basic PCR?). Isolating a gene for production would just be PCR and is pretty easy and robust, you can buy stuff off of eBay to do that (although I'd recommend a better polymerase than taq)
On Sunday, September 18, 2016 at 7:19:00 AM UTC-7, Dean Nunyabis wrote:
I know that Staphylococcus aureus CRISPR is characterized, so what I would do is target thioalcohol to be cut with the CRISPR from Staphylococcus (in which you would supply the Staphylococcus CRISPR on a plasmid)
... [homology] [thioalcohol] [homology]...
\ \ cut^ cut^ \ \
\ \ \ \
-Plasmid-( [homology] [ester] [homology] ) - [sgRNA] - [CRISPR locus] -
The sgRNA, homology, and ester would likely have to by synthesized. That's the easiest way to do that, in my opinion. Sorry if the drawing is bad.
-Koeng
On Sunday, September 18, 2016 at 7:19:00 AM UTC-7, Dean Nunyabis wrote:
Taking a desired trait from one eukaryotic organism and inserting it into another. E.g., isolating the trait that produces thioalcohol in Staphylococcus hominis and replacing it with a trait that produces and Ester.Do I need to be more specific than that or is it going to be a different process every time I want to isolate a trait from one organism in order to insert it into another?Also, how do you decide on a target site for insertion?
On Sunday, September 18, 2016 at 12:08:01 AM UTC-4, Koeng wrote:Those are 2 different objectives. What exactly is your application?-Koeng
On Saturday, September 17, 2016 at 8:07:11 PM UTC-7, Dean Nunyabis wrote:Is there any way to do this on a hobbyist level?
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