Re: [DIYbio] Re: help me setup DIY-BIO home Lab to produce Recombinant Factor VIII

I'm not usually one to try to dissuade people from trying to achieve things others think is impossible, but you should stop and take a step back and look at the scope of your project.

You're trying to make a biologics drug that some company has poured probably hundreds of millions of dollars into, with access to some of the most advanced equipment and best minds in the world, and even at their industrial scale production of this compound, it still costs something like $1.60 per MICROgram. 

If it was as easy as making a plasmid and putting it in E. Coli, it'd have already been done.  I have no idea how it's made but read that they use some weird cell line of a hamster or some other rodent.  Ya, never-mind, the link that I already posted has that information.  There are more steps after it's even excreted by the cells, including some catalyse treatment step and then a separation step.

http://www.rxlist.com/novoseven-drug.htm

On Sun, Jan 18, 2015 at 1:31 AM, Sunil Phani <consults...@gmail.com> wrote:

can some one though light on Gibson assembly approach for a cloning a vector to produce Factor VIII protein

will the size of the DNA to be cloned have any constraints with Gibson assembly approach because the size of Coagulation factor VIII is Size: 2351 amino acids; 267009 Da

On Saturday, January 17, 2015 at 7:31:14 AM UTC+5:30, Sunil Phani wrote:

can someone please help me setup DIY-BIO home Lab to perform mammalian cell culture to produce  Recombinant Factor VIII protein with  mammalian cells(CHO, HKE 293, HEL, etc) and kindly provide me recombinant dna protocols for identification, isolation , extraction, insertion and expansion .my requirement is to prepare pilot project in small scale DIY-BIO lab which can further be scaled.

if possible provide me vectors, cell line and seedstock. i am new to DIY-BIO hence i need helping hands of veterans like you ppl .. your help be highly appreciated

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