Discuss how you can genetically engineer an ecoli cell to...

Discuss how you can genetically engineer an E. coli cell to produce a human protein. Be sure to include in your answer the use of restriction enzymes, ligase; pUC19, X-gal, selective media, and a method for determining that your transgenic E. coli contains your human gene of interest and that it is producing the appropriate protein. Help, please.

Transcript

00:01 In this question we need to discuss the genetic engineer in which we have taken the e.

00:09 Coli cell and we need to transform it in such a way that it starts to produce human protein.

00:19 So we are basically taken a plasmid which is the extra -gromosomal dna.

00:26 So here we have taken puck9.

00:30 As our plasmid.

00:32 This puck 19 is basically the enhanced version of the pbr 322 plasmid and the size of the puck 19 is 2686 base pair long and it also has polylinker sites and it has a ampicillin resistant gene and it can also be selected.

01:00 That is there is insertional activation of the beta galactocidus gene so it basically have the lack z gene here which produces the beta galactosides which breaks the lactose into glucose and galactose and hence that is why if they put it into a xgal medium it will produce a blue color colony but here what we will do is we will insert our gene of interest.

01:35 So let us say this here is our gene of interest and the restriction endonuclease site present on these puck 19 here is of pst and there are many are of eco r1, h1, there are many sites.

02:00 So let us say.

02:01 So let us say.

02:01 Suppose that we have cut this gene of interest with the restriction endonuclease that is sal one and we need to insert it here so that means we will cut this ladsi gene here and here and we will remove this latsy portion and this will be cut with the same restriction endonuclease as our gene of interest now here we have this portion of the gene of interest which will be inserted here so our gene of interest is present here now this lack z gene is basically inactivated so that means lax z gene cannot produce the beta galactosyris and that is why it cannot cut the lactose into the glucose and gallicose so that means the x -gal medium will not be blue in color so that means if a cell is completely transformed it will produce a white column because here the laxygen is deactivated so this is the transgenic e...

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