SBI 4U Genetic Engineering

weedheightswaistBiotechnology

Dec 11, 2012 (4 years and 6 months ago)

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Genetic Engineering is the altering the sequence of DNA molecules




Many new products have been developed through genetic engineering, such as
insulin.




There is also a huge agricultural impact.


o

Frost damage decreased by introducing bacteria that have b
een engineered
to disable the crops from “ice nucleation factor” (which is a protein that is
found on bacterial coats that allows ice to seed and crystallize)




The growth hormone: somatropin is being synthesized.

o

It is a drug that is identical to the hu
man growth hormone: somatotropin.

o

It is used to treat human growth deficiencies that results from a genetic
mutation that causes dwarfism and Turner’s Syndrome.

o

Because it helps build muscle, it is used to
help AIDS patients.

o

There has been some controve
rsy recently
around the availability of somatropin and
athletes. It is a banned substance in sporting
events during international competitions.

o

Another form of the drug: bovine
somatotropin is used in some states to boost
milk production in cows, but it

is banned in
Canada.






The Polymerase Chain Reaction



Polymerase chain reaction: amplification of DNA sequence by repeated cycles of
strand separation and replication.




Before PCR, we could not make numerous copies of the one gene fragment unless
it wa
s inserted into a plasmid.




PCR is a direct method of making copies of a desired DNA sequence.




The process is very similar to DNA replication.


o

Instead of using DNA helicase and DNA
gyrase, PCR uses heat to unwind the DNA
because at high temperatures (94
-
96

C), the
hydrogen bonds break and the strands separate.


o

Next, the two single strands can be used to build
complementary strands


o

Then, in PCR, DNA primers are used because
they are easily produced in the lab (whereas in
replication, we need to put down

RNA primers
first.)


o

We copy the DNA strand by laying down the 5’ to 3’ (starting on the 3’
side of the original DNA)


o

One of the DNA primers is known as the forward primer, and the other is
the reverse primer because they make DNA in opposite directions.


o

The temperature is brought down to 50
-
65

C and it allows for the primers
to stick (anneal) onto the original strands.


o

Once the primers have annealed, the
Taq
polymerase

(DNA polymerase
that can stand high temperatures) can now build the complementary
st
rand.


o

The cycle then repeats itself.


o

There is an exponential increase in the number of targeted DNA.


http://video.google.com/videoplay?docid=6919084111744501197&q=polymerase+chain
+reaction&total=11&start=0&num=10&so=0&type=search&plindex=3




PCR is useful in forensic investigations, medical diagnosis, genetic research




Only requires a small amount of D
NA to work




Used to detect HIV earlier than normal




Used to determine if fossilized specimen were closely related species.


Restriction Fragment Length Polymorphism




Polymorphism: any difference in DNA sequence, coding or non
-
coding, that can
be detected b
etween individuals




All individuals from the same species are said to be polymorphic unless they are
identical twins.




Polymorphisms happen in both coding and non
-
coding regions.




In coding areas, polymorphisms may be used to
identify specific mutations:
for example, a person
who carries the allele for sickle cell anemia carries a
different allele for


globin that gives red blood cell a
sickle shape compared to a person who does not have
the disease.




We can have polymorphisms in our non
-
coding
regions to
o, and are exploited in forensic
investigations.




Restriction fragment length polymorphism
(RFLP) analysis

is a technique where DNA regions
are digested using restriction endonucleases and
subjected to radioactive complementary DNA probes
to compare the di
fferences in DNA fragment lengths
between individuals.


o

the patterns that result after running these cut
DNA’s through a gel are usually represented
by a big smear because there is so much DNA
available it doesn’t separate well.

o

so we have to a procedure t
hat allows the
DNA in an electrophoresis gel to be
transferred to a nylon membrane while
maintaining the position of the DNA band
fragments: called
Southern blotting



we denature the DNA so it becomes
single
-
stranded and the negatively
charged DNA will stic
k to the blotting
nylon membrane.



The membrane is then immersed in a
solution that has radioactive
complementary nucleotide probes for
specifically chosen areas on the DNA



Nylon membrane is then placed against
x
-
ray film and the radioactive probes cause ex
posure on the film,
to see a pattern.


p. 298 # 1, 3, 4, 5

p. 300 # 8, 9.