Comment · Fri, August 21, 2015 · Ceretropic
Anyone else interested in P21 nasal spray from Ceretropic?
Original post in this thread
SimianSound · 18 points
I'm hoping this post serves as a petition of sorts to the awesome vendors over at Ceretropic. Show your support if you're also waiting on this!
What they were answering
SangersSequence · 8 points
They use Invitrogen's one-step seamless cloning and assembly reaction to insert the coding sequence for the peptide between a stop sequence and a highly specific protease cleavage site (downstream of a chaperone which is unnecessary since you aren't folding) in the payload plasmid for the transfection virus.
For P21, like most peptides of this incredibly small size you'd be able to insert the sequence using a block of 60-75 nucleotides including the overlapping regions for seamless cloning. Essentially a sense and antisense duplex primer, which is actually how we ordered the small ones from IDT, you can get much large proteins using gBlocks.
Since none of your peptides require disulfide bonding, it might actually be possible to produce them using the e. coli plasmid system that doesn't require a lentiviral transfection. Although you'd have to be sure to use an endotoxin free e coli strain (like this one: http://www.immunosource.com/media/cleancoli_poster_22-apr-2013.pdf)
You are only able to use standard amino acids, unless you want to get into using additionally modified e coli strains (amber codon supression: http://www.ncbi.nlm.nih.gov/pmc/articles/PMC3982533/) in which case you could add any novel amino acid you're willing to make a t-rna for.
This part isn't exactly my field, I'm studying gene mutations that cause defects in developmental neurogenesis for my PhD (whi…
u/MisterYouAreSoDumb · Ceretropic
That's seems very interesting! I'll have to look into it a bit. Is Lucigen the people to speak to about it?