1. DNA Technology
05/25/12 cott

2. What makes one allele different
from another allele?
 All alleles for a particular gene contain DNA
instructions for the synthesis of the same
protein
 BUT…slight differences in the base
sequences of two alleles, such as:
 AGGCTTAGA, vs.
 AGGCTAAGA
 Can cause slight differences in the protein produced
by the alleles
05/25/12 cott

3. Example:
 TRAIT: BLOOD CLOTTING
 GENE: CLOTTING FACTOR
 LOCUS X CHROMOSOME
 ALLELES: ACGGTACT (XH - normal)
ACGATACT (Xh - hemophilia)
05/25/12 cott

4. How is the genetic code
translated into protein structure?
 Proteins are chains made of many amino acids
linked together
 The sequence of amino acids is specified by the
base sequence of the DNA…each sequence of 3
bases codes for a different amino acid
GAC ACA CAG GGG AAG DNA
Chain of amino
acids
05/25/12 cott

5. DNA Technology: Genetic
Engineering
 Scientists use knowledge of structure of
DNA to study and change DNA molecules
 Uses tools:
 DNA extraction – removing DNA
 Restriction enzymes – cutting DNA
 Gel electrophoresis – separating DNA
 PCR
 DNA Fingerprinting
05/25/12 cott

6. DNA Extraction
 Place cells in
detergent to break
down membranes
 Place cell extract in
ethanol
 DNA is insoluble in
ethanol, so it comes
out of solution, and
can be removed for
study
05/25/12 cott

7. Restriction Enzymes – Cutting
DNA
 Enzymes that recognize a particular short
DNA sequence, and then cut the DNA
strand within that sequence.
 1st discovered in bacteria which use the
enzymes to cut and destroy viral DNA.
 DNA molecules are too large to be
analyzed as a whole.
05/25/12 cott

8. Restriction Enzymes
 Eco RI – restriction enzyme found in E.
Coli.
 EACH RESTRICTION ENZYME
RECOGNIZES A DIFFERENT SEQUENCE
OF DNA AND WILL CUT THAT
SEQUENCE ONLY!!!
 DNA Restriction - a MAD GOOD ANN'Y!
05/25/12 cott

9. Restriction Enzymes
Staggered Cut
Staggered (Eco Ri) cut
5’ - GAATTC – 3’
3’ - CTTAAG – 5
to produce sticky ends:
5’ G AATTC – 3’
3’ CTTAA G – 5’
05/25/12 cott

10. Restriction Enzymes
Blunt Cut
Blunt Cut:
5’ – GTTAAC – 3’
3’ – CAATTG – 5’
Blunt Ends:
5’ GTT AAC – 3’
3’ CAA TTG – 5’
Blunt ends can be attached to any other DNA
that produces blunt ends.
05/25/12 cott

11. Restriction Enzymes
Example:
Enzyme called
EcoR I
Cuts DNA
anywhere it
finds sequence
TTAA
05/25/12 cott

12. Gel Electrophoresis
 Means of separating, by size, the
DNA fragments produced by
restriction enzyme cuts.
 Compares genes of different
individuals or organisms
05/25/12 cott

13. Gel Electrophoresis
1. DNA fragments placed into wells in gel slab
2. Electric voltage is applied to gel.
3. DNA (negatively charged) migrates to (+) end of gel.
4.05/25/12
Smaller the fragment, faster and farther it moves.
cott

14. Cell Transformation
 What happens during cell
transformation?
 How can you tell if a transformation
experiment has been successful?
05/25/12 cott

15. Plasmid
 Small DNA molecules found naturally in
bacteria.
 Useful for DNA transfer. WHY?
 It has a DNA sequence.
 If a plasmid containing foreign DNA gets
into a bacterial cell >>>>> replication
occurs.
05/25/12 cott

16. Recombinant DNA
 Plasmids have
genetic markers
 Make it possible
to distinguish
foreign DNA
05/25/12 cott

17. Recombinant DNA
 If transformation is successful DNA
is combined (recombinant) into on
of the chromosomes of the cell.
 Applications for genetic
engineering:
 Create human forms of protein >>>
insulin, HGH, clotting factor.
 Cloning
05/25/12 cott

18. 05/25/12 cott

19. DNA Fingerprinting
A practical use of DNA technology
05/25/12 cott

20. DNA Fingerprinting
 Just as no two people have the
same fingerprint, no two people
(except identical twins) have same
DNA.
05/25/12 cott

21. DNA Fingerprinting
 Differences between individuals
due to different sequences of
nucleotide bases
 Remember, different alleles are result
of differences in base sequences
 Ex: AACTGGCA vs.
AACCGGCA
05/25/12 cott

22. DNA Fingerprinting
 In addition, between genes,
chromosomes contain large
amounts of DNA repeats
 Do NOT code for proteins
 Example: AAAATTTTAAAATTTT, etc.
 Number of repeats between genes
varies from person to person
05/25/12 cott