Genetic engineering
Recombinant DNA technology
Questions:
• Name 3 things you know about bacteria.
• What are some characteristics that make
bacteria a good choice for genetic engineering?
• Why are we using bacteria for biotechnology
rather than some other organism?
Bacteria are useful hosts.
1.
2.
3.
4.
They are easily grown
They are cheap to grow
They grow fast
They are easily manipulated in the laboratory
1. DNA can be inserted - transformation
2. DNA can be easily isolated
5. Bacteria contain natural plasmids and viruses
which are useful vectors for recombinant DNA
Bacterial
plasmids
•
Most bacterial
DNA is on a single
large
chromosome, but
some DNA is in a
small circle called
a plasmid.
Bacterial Plasmids in Nature
1.
2.
Occur naturally in bacteria
and usually carry genes
that are useful but not
essential to survival: e.g.
genes which make
bacteria resistant to
antibiotics.
Plasmids are released by
dead bacteria and
absorbed by those still
living thus genetic
information is exchanged
(sexual reproduction?).
Bacterial Plasmids in Nature
3. Some plasmids even
contain genes that
build a transfer tube
between bacteria.
4. There can be as
many as several
hundred copies of a
single plasmid in
each bacteria.
Plasmid Structure
1. Plasmids only need an
“origin or replication” and a
“useful” gene to be
considered complete.
2. Molecular biologists have
been able to “insert” custom
built restriction sites into
many plasmids so they can
be used to “insert” DNA
fragments from other genes
into them and thus have a
way to propagate those DNA
pieces.
Plasmids can be manipulated
easily in the laboratory
1.
2.
3.
4.
Plasmids can be collected from bacteria.
Restriction enzymes can be purified and used to identify and cut
out specific sequences of DNA along with the plasmid vector.
Ligase (enzyme) can be purified and used to “glue” pieces of
DNA together.
Bacteria can be transformed by taking in plasmids given to
them.
Transforming Bacteria
• When a bacterial cell
takes in a plasmid from
the environment, it has
new DNA (and
therefore, new traits).
• Scientists say that the
bacteria has been
transformed.
How to produce human insulin from
bacteria and become a multimillionaire
1.
2.
3.
4.
5.
6.
Isolate plasmid DNA from
bacteria and insulin gene
from human.
Cut both DNAs with the
same restriction enzyme.
Mix the DNA together with
ligase.
Insert the new DNA into
bacteria (transformation).
Use antibiotics to kill any
bacteria without the
plasmid.
Grow bacteria and harvest
the insulin.
Human DNA
Insulin gene
Insulin
Purification
Some Links
• Basic construction of recombinant DNA
http://www.dnai.org/b/index.html
– Click on “techniques” then “cutting and
pasting”
• An excuse to study on Youtube!!
http://www.youtube.com/watch?v=x2jUMG
2E-ic
Your Turn
• Plasmid Maps
–Restriction Enzyme Challenge
–Activity I and II
• Recombinant DNA
–Paper plasmid Activity