Download Recombinant DNA Technology Biotechnology

Biotechnology
Recombinant
DNA
Technology
7/7/2017
Bellwork

Explain what genetic
engineering means to you?
7/7/2017
Objectives
Define recombinant DNA
 Create a paper model of a
recombinant plasmid and identify the
key components

7/7/2017
Key Terminology
Recombinant DNA molecule
 Chimera
 DNA ligase
 Plasmid
 Vector
 Transformation
 Chromosomal DNA

Overview

Recombinant DNA Technology involves
using methods to create new DNA
molecules

New DNA molecules made by piecing
together DNA from different organisms
 This recombinant DNA (rDNA) is known as a
chimera
Goat boy
Examples of Recombinant DNA
Technology
rhInsulin, marketed by Eli, Lilly and
Company
 Recombinant human growth hormone
(Nutropin, marketed by Genentech, Inc)
 Recombinant “activase” (marketed
Genentech, Inc)-dissolves blockages in
blood vessels

Significance of rDNA
These proteins normally are produced in
very small amounts or are synthesized at
the wrong time in the human body
 By being able to make these products in
large amounts in other organisms, like
bacteria.

 Can
be used to help individuals that are
deficient in these proteins
Plasmid DNA


A small circular double
stranded DNA found in
bacteria and yeast
Separate from the
chromosomal DNA

Chromosomal DNA controls growth and
division of organism
Characteristics of Plasmid DNA



Not involved in regulating growth and division of
organism.
Consider autonomous in that it has control over
its replication
Presence of plasmids in bacteria often present
to protect against humans medicines by carrying
antibiotic resistance genes
 Prevents antibiotics from killing the bacteria
Why is a Plasmid called a
Vector ?

Because the plasmid can replicate and
carry recombinant DNA into organism of
interest
Making a Recombinant DNA
Molecule

Step 1: Isolation of human DNA encoding
Insulin.
 Isolate
Human DNA
 Cut DNA into fragments with restriction
enzymes
 Identify the DNA fragment that encodes for
the protein, insulin
Making a Recombinant DNA
Molecule

Step 2: Place DNA molecule into a
cloning vector (commonly plasmid DNA)
 Cut
plasmid with same restriction enzyme
used to make DNA fragments that contain
insulin
 Ligate fragments into the plasmid with DNA
ligase
 DNA ligase is an enzyme that “glues” the
DNA fragments together to create the
recombinant DNA molecule
Picture of Cloning Vector
Transforming Bacteria with
Recombinant DNA

Step 3: The recombinant DNA molecule is
taken up by the Bacterium by
transformation

When a bacterial cell takes up foreign DNA
(e.g. plasmid vector containing human
insulin), it is said to be transformed
 Bacteria then allowed to reproduce to make
lots of cells containing the recombinant DNA
molecule
Think-Pair-Share
Why is the plasmid with the
human insulin DNA
considered to be a
recombinant DNA
molecule or chimera?
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Identify the Bacteria with the
Recombinant DNA molecule
Must eliminate bacteria cells that do not
carry the recombinant plasmid
 Accomplished by growing bacteria on
media containing an antibiotic

–chemicals that prevent the growth
and survival of bacteria
 Antibiotic

How does the antibiotic work?
Antibiotic Resistance
Bacterium Takes Up Cloning
Vector

Bacterium takes up cloning vector will now
become ampicillin resistance.
 In
other words, will not be killed by the
presence of ampicillin in the growth media
 The bacteria without the cloning vector will be
killed because do not contain the ampicillin
resistant gene.
Think-Pair-Share
You have transformed bacteria with the
plasmid containing the human insulin
gene. Only 10% of the bacteria take up
the recombinant DNA molecule. Predit
which petri plate will have more bacteria
growth, the one + ampicillin or the one –
ampicillin. Explain your reasoning.
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In 12 words or less, list key
concepts you learned today
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Q & A: Recombinant DNA
1)
2)
3)
4)
5)
Why is it useful to create bacteria that contain a new
gene?
What is the purpose of restriction enzymes in this
process?
What is the purpose of DNA Ligase?
Describe the 3 steps for making recombinant DNA.
Explain the specific steps you would use to create
bacteria with a human growth factor gene.