Biotechnology
AP Biology 2015 - 2016
Campbell Biology in Focus: Chapter 13 Section 4
I. Definitions
A. Genetic Engineering: process of manipulating genes and genomes
B. Biotechnology: process of manipulating organisms or their components for the purpose of
making useful products.
C. Recombinant DNA: DNA that has been artificially made, using DNA from different
sources. eg. Human gene inserted into E.coli
D. Gene cloning: process by which scientists can product multiple copies of specific
segments of DNA that they can then work with in the lab
II. Tools of Genetic Engineering
A. Restriction enzymes (restriction
endonucleases): used to cut strands of DNA at
specific locations (restriction sites)
B. Restriction Fragments: have at least 1 sticky
end (single-stranded end)
C. DNA ligase: joins DNA fragments
D. Cloning vector: carries the DNA sequence to
be cloned (eg. bacterial plasmid)
III. Gene Cloning
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IV. Applications of Gene Cloning
V. Polymerase Chain Reaction - technique that produces many copies of a DNA molecule
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VI. Techniques to track or follow DNA
A. Probe hybridization: find and track gene of interest
B. Gel electrophoresis: separate DNA molecules by size and charge
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VII. DNA Sequencing: general process is similar to PCR
Primer design:
Ingredients:
1. target DNA to be sequenced
2. primer - short strand of DNA that will
hybridize to one strand of the DNA to be
sequenced
Sequencing plasmid:
(target DNA will be inserted into polylinker)
Usually the targeted DNA has been inserted into
a known plasmid, so the plasmid DNA is used
to design the primer so that replication soon
enter unknown DNA sequence
3. polymerase
4. regular nucleotides
Normal nucleotide:
Dideoxynucleotide:
5. a small amount of dideoxynucleotides that
are labelled with a fluorescent tag
(dideoxynucleotides lack a 3' OH group so a
polymerase cannot continue after inserting a
dideoxynucleotide)
Procedure:
1. Allow the polymerase to replicate the target
DNA to be sequenced.
2. Every time a polymerase uses a
dideoxynucleotide (ddC, ddG, ddA or ddT)
then that replication process will stop
3. The result is a mixture of DNA molecules
that are basically identical but have different
lengths and end with a fluorescent marker
4. Separate the mixture of DNA through gel
electrophoresis
5. The flourescent tags are read by computer as
they exit the gel
6. 500 to 1000 bases of DNA can be read.
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Typical Output from DNA Sequencing:
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