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BIOTECHNOLOGY
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What is biotechnology?
• Aspect of technology that uses:- biological data- molecules - organisms for alternative practices
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Bioremediation
• The use of living microorganisms to transform harmful substance into non-toxic compounds
• Example:• EXXON VALDEZ oil spill in March 1989
– 50,000,000L of oil spewed into the Alaskan Sea– Covering 3,400km2
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Exxon Valdez Clean-up
• Used:– Physical barriers– Pumps– *microrobes
• Scientists released microbes with oil degrading enzymes – Research is now looking into adding nutrients (O2)
to the oil spill to increase microbe growth
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Uses of Biotechnology1. Investigating genetic disorders
2. Altering genetic make-up of organisms- production of useful proteins
3. Analyze DNA evidence
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Biotechnology Tools• Like with any trade, there are certain tools
needed to complete a specific task
• Biotechnologists, or molecular biologists, use biological molecules– to cut, join, & replicate DNA
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Biotechnology Tools1. Restriction Endonucleases / enzymes2. Methylases3. DNA ligase4. Gel Electrophoresis5. Plasmids6. Transformation7. PCR8. RFLPs9. DNA sequencing
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1. Restriction Endonucleases (RE’s)
AKA Restriction Enzymes (RE’s)• Enzymes that are able to cleave (cut) double
stranded DNA into fragments – Only cut at specific base pairs
• Each RE has its own recognition site– Specific DNA sequence – 4-8 base pairs in length– Characterized by a palindromic sequence
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1. Restriction Endonucleases (RE’s)
Ex. EcoRI is a restriction enzymeCuts DNA when it sees the following sequence 5’ – G A A T T C – 3’3’ – C T T A A G – 5’
5’ – G A A T T C – 3’3’ – C T T A A G – 5’
5’ – G A A T T C – 3’3’ – C T T A A G – 5’
**Considered palindromic because both strands have the same sequence when read 5’ 3’
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1. Restriction Endonucleases (RE’s)
How RE’s work (text form)1.Scan DNA for cognition site2.Once found, it binds and uses a hydrolysis rxn
to break phosphodiester linkages between A and G nucleotides on each strand
3.H-bonding between nucleotides is distrupted4.Two fragments are produced
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1. Restriction Endonucleases (RE’s)
The ‘ends’ produced by RE’s depend on what RE is used.
There are two types of ‘ends’:1.Sticky Ends2.Blunt Ends
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1. Restriction Endonucleases (RE’s)
Sticky Ends• fragment end has short SS DNA with
unbound base pairs• Generally more useful
– Easier to join to other sticky ends
• Ex. EcoRI 5’ – G A A T T C – 3’3’ – C T T A A G – 5’
5’ – G A A T T C – 3’3’ – C T T A A G – 5’
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1. Restriction Endonucleases (RE’s)
Blunt Ends• Fragment ends are fully base paired• Less useful
– as harder to bind to other blunt ends
• Ex. SmaI5’ – G G G C C C – 3’3’ – C C C G G G – 5’
5’ – G G G C C C – 3’3’ – C C C G G G – 5’
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1. Restriction Endonucleases (RE’s)Question:Why would it be better to have 4-8 base pairs in
your recognition site than 2 base pairs?
Answer: The shorter the recognition sequence, the more
likely the RE is to cut & may disrupt a gene
*probability of finding a 6bp recognition site:4 x 4 x 4 x 4 x 4 x 4 = 4,096 1 in 4,096 bases.