Download Biotechnology, Part I

Biotechnology:
The Basics
Honors Biology
Honors Biology
The Future
General Biology
Biotechnology
 The genetic manipulation of organisms

Humans have been doing this for
thousands of years

General Biology
Selective Breeding - Plant & animal breeding
for specific traits
Breeding of food plants
 “Descendants” of the wild mustard

Brassica spp.
General Biology
Breeding of food plants
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Animal husbandry/breeding
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Bioengineering Tool Kit
 Basic Tools

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Restriction enzymes
Ligase
Plasmids/cloning
 Advanced Tools

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PCR
DNA Sequencing
Gel electrophoresis
Southern Blot
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Biotechnology
 Genetic Engineering: The process of
manipulating genes for practical
purposes.

Faster and more reliable than selective
breeding.

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Genetic Engineering may involve Recombinant
DNA Technology.
Recombinant DNA
 Recombinant DNA – DNA from two
organisms put together in 1 organism.
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Plants and animals that contain recombinant
DNA are known as Transgenic Organisms
Genetic Engineering: Steps


Let’s look at how bacteria
can make human insulin!
4-Step process.
1. Cutting DNA
2. Making recombinant DNA
3. Cloning
4. Screening
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Step 1: Cutting DNA
 First, you have to cut out the gene your
want.
 Cutting DNA is done by restriction
enzymes (molecular scissors!)


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Enzymes that cut at specific nucleotide
sequences using palindromes
Produces “sticky ends”
What do you notice about these
phrases?
radar
palindromes
racecar
Madam I’m Adam
Able was I ere I saw Elba
a man, a plan, a canal, Panama
Was it a bar or a bat I saw?
go hang a salami I’m a lasagna hog
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Restriction enzymes Madam I’m Adam
cut DNA at specific sequences

restriction site
symmetrical “palindrome”
 produces protruding ends
CTGAATTCCG
GACTTAAGGC

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
 Action of enzyme



sticky ends
will bind to any complementary DNA
CTG|AATTCCG
GACTTAA|GGC
 Many different enzymes

named after organism they are found in

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EcoRI, HindIII, BamHI, SmaI
Restriction Enzymes
 Cut DNA at specific sites

leave “sticky ends”
restriction enzyme cut site
GTAACGAATTCACGCTT
CATTGCTTAAGTGCGAA
restriction enzyme cut site
GTAACG AATTCACGCTT
CATTGCTTAA GTGCGAA
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Sticky ends
 Cut other DNA with same enzymes


leave “sticky ends” on both
can glue DNA together at “sticky ends”
GTAACG AATTCACGCTT
CATTGCTTAA GTGCGAA
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gene
you want
GGACCTG AATTCCGGATA
CCTGGACTTAA GGCCTAT
chromosome
want to add
gene to
GGACCTG AATTCACGCTT
CCTGGACTTAA GTGCGAA
combined
DNA
Sticky ends help glue genes
together
cut sites
gene you want
cut sites
TTGTAACGAATTCTACGAATGGTTACATCGCCGAATTCACGCTT
AACATTGCTTAAGATGCTTACCAATGTAGCGGCTTAAGTGCGAA
AATTCTACGAATGGTTACATCGCCG
GATGCTTACCAATGTAGCGGCTTAA
sticky ends
cut sites
isolated gene
chromosome want to add gene to
AATGGTTACTTGTAACG AATTCTACGATCGCCGATTCAACGCTT
TTACCAATGAACATTGCTTAA GATGCTAGCGGCTAAGTTGCGAA
DNA ligase joins the strands
sticky ends stick together
Recombinant DNA molecule
chromosome with new gene added
TAACGAATTCTACGAATGGTTACATCGCCGAATTCTACGATC
CATTGCTTAAGATGCTTACCAATGTAGCGGCTTAAGATGCTAGC
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Step 1: Cutting DNA
 You also need to cut a vector.

Vector - agent that will carry the gene we
want into another cell.
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Common vectors are bacterial plasmids, yeast, or
viruses
What is a Plasmids?
 Small, supplemental circles of DNA in
bacteria
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Self-replicating
Carry extra genes

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Genes for antibacterial resistance
Can be exchanged between bacteria
Can be imported from environment
(transformation!)
General Biology
Step 2: Making Recombinant
DNA
 The gene of interest is pasted into the
vector DNA along with a gene for
resistance to a specific antibiotic.

Ligase is used as glue!

In our example, the human gene for insulin
production is combined with a plasmid.
Step 3: Cloning
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Bacteria then take up (transformation!)
the recombinant plasmids.
In a process called gene cloning,
many copies of the gene of interest
are made each time the host cell
(bacteria) divides.
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Step 4: Screening
 Bacteria cells that have the gene of
interest must be separated from cells
that do not

Cells are grown on antibiotic plates - only
those with the gene of interest and
antibiotic resistance survive!
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transformed
Grow bacteria…make more
bacteria
gene from
other organism
recombinant
plasmid
+
vector
plasmid
grow
bacteria
harvest (purify)
protein
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Uses of Genetic Engineering
 Genetically Modified Organisms (GMO)

“Fishberries”, Golden Rice,“Wool off”
gene in sheep,“Roundup-ready” bacterial
gene in soybeans, BT Corn
 Health: Drugs, vaccines, Gene Therapy
 Cloning: Farm animals, pets, humans?
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Cut, Paste, Copy, Find…
 Word processing metaphor…

cut
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paste
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restriction enzymes
ligase
copy

plasmids
– bacterial transformation
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is there an easier way??
find
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Honors Biology
????