Genetic Engineering

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16 Δεκ 2012 (πριν από 4 χρόνια και 4 μήνες)

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Genetic
Engineering

What is Genetic Engineering?


Genetic Engineering = inserting a foreign
gene of interest into a host to transcribe and
translate a particular protein.


Ex. Inserting the human insulin gene into
bacteria to mass produce it.

Image taken without permission from http://www.medicalprogress.org/uploads/images/insulin%20inject%20WSU%20210.jpg

General Steps


Obtain the gene of interest (ex. insulin
gene)


Insert the gene into the host (ex. bacteria)


Allow the host to multiply and express the
foreign gene


get your desired protein!


Get lots of cells that can make the protein =
clones


The Big Picture


The inserted gene is transcribed and
translated using the RNA Polymerase,
ribosomes and other resources in the cell

Plasmids


Circular DNA


Extrachromosomal


NOT part of the
E. coli

genome


“extra” DNA


Contain a few non
-
essential genes


Can give the bacteria additional “traits”


Depends on the genes on the plasmid


Can be exchanged between bacteria

chromosomal DNA

plasmids

Recombinant plasmids


Plasmids can be modified in biological labs


Modified plasmid = Recombinant plasmid


Plasmids can be used as cloning vectors to
get the recombinant plasmid into
E. coli


Cloning vectors = way to get the gene of
interest into the host

Transformation


Process in which
foreign DNA is
physically inserted into
host
E. coli

cells.


E. coli

that contains
recombinant plasmid =
Transformed cell


Image taken with out permission from
http://summerschool.at/static/irismaria.schoenbrunner/imag
es/transformation.png


Transformation Steps


Recombinant plasmids
and host E. coli are mixed
together


CaCl
2

is added


The Ca
2+

ions neutralize
the negative charges on
plasmid DNA


Help plasmid enter the
membrane

Image taken without permission from

Transformation Animation. Available at
http://www.dnai.org/b/index.html


Transformation Steps


Heat Shock


By rapidly changing the temperature of the
solution, temporary pores are opened in the
membrane


Creates an opening for the plasmids to enter the
E. coli

Transformation


Transformation is not 100% successful


After transformation


Some cells will contain plasmid =
transformed


Some cells won’t contain plasmid =
untransformed


In a later step, you will determine which
cells were transformed

E. Coli

as a host


E. coli is a good host because:


Reproduce quickly (once every 20 minutes)


Nonpathogenic (the strain we use is not
harmful)


Genome fully characterized (all genes have
been sequenced)