Genetic Engineering I
Restriction Enzymes and Gel
Electrophoresis
How are changes made to DNA
• Genetic engineering: Scientists use their
knowledge of the structure of DNA and its
chemical properties to study and change DNA
molecules
• Different techniques are used to:
- Cut DNA into smaller pieces
- Identify the sequence of bases in a DNA molecule
- extract DNA from cells
- Make unlimited copies of DNA
The tools of molecular biology
• Cutting DNA: restriction enzymes are used to
cut DNA into smaller pieces
• Separating different length of DNA: gel
electrophoresis is used to separate different
fragments of DNA by their size
Restriction Enzymes
• Naturally found in different types of bacteria:
- Bacteria use restriction enzymes to protect
themselves from foreign DNA injected by
viruses
• Function: Restriction enzymes cut DNA at
specific sequences, they are also called
“ molecular scissors”
Restriction Enzymes
• There are 2 types of bonds found in DNA:
1. Covalent bonds : found inbetween
nucleotides within a single strand
2. Hydrogen bonds: found inbetween the 2
strands
Restriction Enzymes
• Restriction enzymes cut the covalent bonds of
DNA molecules at specific sequences called
restriction site:
Do you notice anything interesting
about the sequence of the two strands?
• 5’ GAATTC 3’
3’ CTTAAG 5’
• Remember: DNA is composed of anti-parallel
strands
• Palindrome sequence: a sequence that is the
same whether read 5’ to 3’ on one strand or 5’
to 3’ on the complementary strand
• Some English palindromes: Dad, racecar,
nurses run
Restriction Enzymes
• When restriction enzymes cut they produce
either:
- Sticky ends:
- Blunt ends:
Gel electrophoresis
• Gel electrophoresis separates molecules on the
basis of their charge and size:
- DNA molecules are negatively charged
- Different sized DNA fragments are placed in a gel
made of agarose (a sugar found in seaweed)
- Electricity is applied to the gel
- DNA fragments will move across the gel at a
speed dependent on their size:
the longer DNA fragments will move slower
the shorter DNA fragments will move faster
Gel electrophoresis
Wells
Gel electrophoresis
Applications for restriction enzymes
and gel electrophoresis
1. Evolutionary study: Comparing DNA samples
from different organisms to measure
evolutionary relationships
turtle snake rat squirrel
–
DNA

+
1
2
3
4
5
1
2
3
4
fruitfly
5
Applications for restriction enzymes
and gel electrophoresis
2. Medical diagnostic: Comparing normal alleles
to disease alleles:
e.g. test for Huntington's disease
–
DNA
chromosome
with normal
allele 1
chromosome with
disease-causing
allele 2

+
Application for restriction enzyme and
gel electrophoresis
3. Forensics: comparing DNA samples from a
crime scene with suspects & victim
suspects
S1 S2 S3
crime
scene
V sample
–DNA

+
Application for restriction enzyme and
gel electrophoresis
4. Paternity: Who’s the father?
–
DNA

+
Mom
F1
F2
child