* Why do we want to make Eukaryotic cell proteins?
* Because when eukaryotic proteins are expressed in prokaryotic cells problems like been unstable, no biological activity and prokaryotic contaminants ( pyrogens) can occur. * Also to make the protein as natural as possible
* The same expression vectors as we need for pro
* We don’t need shangdolron coz that only for pro
Transformation: in Eukaryotic cells: in humans means that difference in growth characters of cells, for example, the cells get transformed to cancer cells Transfection: uptake of foreing DNA into eukaryotic cells resulting in inherited change, the same thing as we did in E.coli transformation Why would we want to use yeast as an expression system
* We know a lot about it
* Its easy to deal with
* Strong promoters
* GRAS= means generally known as safe organisms, we put them in bread, wine etc * Most known safe yeast is saccharomyces cerevisiae
Three types of yeast expression vectors
* Yep: 30 copies but not stable
* Yip: only one copy but very stable
We have inducible and constitutive promoters
A comparsion between saccharomyces and p.pastores, p.pastores has the following advantage * High biomass yield
* Strong promoters
* Efficient secretion
Fungi is also been used for protein expression system
Yeast may not have the correct post –translational modifications so we have to use the closer organisms to achive what we want, for example we use insect cells, mouse cell or human cells Generalized mammalian expression vector
* We still have to do the shuttle plasmid, an E.coli gene and mammalian Why would have an introns increase expression?
* If it does not have introns then it wont be expressed as much as the onces that have introns * The agents are going to kill the Euk by different actions * A marker Gene prevents the agents from doing the actions that would have lead to Euk death What do we...