Genetic Engineering Ppt

Genetic EngineeringGenetic Engineering

First, the nucleus of human cells are burst

Human cellNucleus


The chromosomes are cut up into small fragments and the required gene identified.

Chromosome fragments

Fragment containing required gene


Next the fragments are spread out and the required one isolated.

Segment with required gene


Cytoplasm

Bacterial chromosomeBacterial cell wall

Plasmid

Structure of a typical bacterium


Plasmid

Plasmids are loops of DNA separate from the main chromosome. They carry genes for things like antibiotic resistance. This makes them very useful to theGenetic engineer.


In the above plasmid, the YELLOW gene is one that gives the bacterium resistance to one antibiotic (eg Penicillin).

PP

TT

The GREEN gene gives resistance to a different antibiotic (eg Tetracycline)


By using special enzymes, we can make a cut in the midst of ONE of theseantibiotic resistance genes.In this example, we will cut open the ‘T’ gene

PP

TT

Cut here


Next, we introduce the prepared HUMAN gene to the mixture. If all goes according to plan, the human gene will fit into the cut in the plasmidso that the green ‘T’ gene will no longer work correctly.

Prepared human gene


As plasmids are extremely small, we cannot tell by looking which ones have gotthe human gene in the right place. We need to use a ‘shotgun’ approach andincubate thousands of plasmids with hundreds of bacterial cells

No P or T gene

Intact P gene and Intact P gene and ‘‘defectivedefective’’ T gene T gene

P and T Genes intact


Some cells will take up the recombinant plasmid, some will take up original plasmids, others will take up no plasmds at all or ones without antibioticresistance genes.

Required cellRequired cell Cell with P and T intactCell with P and T intact Cell with neither P or Cell with neither P or TT


An agar plate containing Penicillin is used to allow only those cells which havetaken up a suitable plasmid to survive and divide. These cells must have resistanceto Penicillin

Agar containingpenicillin

Colonies growing from single cells that are resistant to penicillin


Next, these colonies are sub-cultured onto agar containing tetracycline. Only cells resistant to BOTH antibiotics will be able to grow. We are interested in those cells which WON’T grow in the presence of Tetracycline


Next, these colonies are sub-cultured onto agar containing tetracycline.

These cells must have intact T genes

These cells must have intact P genes and defective T genes


This colony will probably have the correct plasmid to produce the product from thehuman gene. Cells from this colony will be grown on a large scale and the mediumanalysed for the presence of the product from the human gene, eg growth hormone

Genetic Engineering Ppt

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