Recombinant Recombinant DNA DNA

TechnologyTechnology

Pre-AP Biology

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BellworkBellwork Explain what genetic

engineering means to you.

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Key Terminology Recombinant Chimera Plasmid Cloning Vector Ligate / DNA Ligase Transformation Transgenic

ObjectivesObjectives

Define recombinant DNA Explain how bacteria are genetically

engineered Create a paper model of a recombinant

plasmid and identify the key components Describe examples of Biotechnology in

plants and animals

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Overview

Recombinant DNA Technology creates new DNA molecules made by piecing together DNA from different

organismsThis recombinant DNA (rDNA) is also 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”

dissolves blockages in blood vesselsmarketed by Genentech, Inc

Significance of rDNA

These proteins normally are produced in very small amounts in animals

Making these proteins in large amounts in other organisms, like bacteriaCan be supplemented in individuals that are

deficient in a protein, like InsulinCan be made cheaper and safer than other

methods

Making a Recombinant DNA Molecule Step 1: Isolation of human DNA encoding

Insulin: Isolate Human DNACut DNA into fragments with a restriction

enzyme Identify the DNA fragment that encodes for

the protein Insulin

Making a Recombinant DNA Molecule

Step 2: Place DNA molecule into a cloning vectorCloning vector is commonly a plasmid

small circular piece of DNA found in bacteria

separate from its chromosomal DNA A plasmid is called a vector

when used to carry the desired DNA into the organism

Making a Recombinant DNA Molecule (Step 2 continued)Fragments of DNA are put into the plasmidCut plasmid with same restriction enzyme used to

make DNA fragments Ligate fragments into the plasmid with DNA ligase

DNA ligase “glues” the DNA fragments together

TThinkhink-P-Pairair-S-SharehareWhy is the plasmid with the

human insulin DNA considered a recombinant DNA molecule or chimera?

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Transforming Bacteria with Recombinant DNA Step 3: The recombinant DNA molecule is

taken up by the Bacteria Bacteria are said to be transformed

When they take up a foreign gene in a plasmid e.g. when they take up the human insulin gene

Transformation Process by which bacterium takes up and

expresses foreign DNA Happens in nature when bacteria die and

release their DNA, and other bacteria take up the DNA

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TThinkhink-P-Pairair-S-SharehareIYOW define transformation

of bacteria.

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Identify the Bacteria with the Recombinant DNA molecule Not all bacteria will be transformed Must eliminate bacteria that do not carry the

recombinant plasmid Accomplished by growing bacteria on media

containing an antibioticAntibiotic –chemicals that prevent the growth

and survival of bacteria How does the antibiotic work?

Definition

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Antibiotic - “against life” substance that kills bacteria or

prevents them from replicating

Cloning Vector

The plasmid that is used as vector is also given an antibiotic resistance gene

The vector shown has the resistant gene to ampicillin

How Bacteria Acquire Antibiotic Resistance

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Bacteria Take Up Cloning Vector

Bacteria that take up cloning vector will also be ampicillin resistant In other words, transformed bacteria will not

be killed when grown on agar containing ampicillin

The bacteria without the cloning vector will be killed because they do not contain the ampicillin resistance gene

TThinkhink-P-Pairair-S-SharehareYou have transformed bacteria with the

plasmid containing the human insulin gene. Only 10% of the bacteria take up the recombinant DNA molecule. Predict which petri plate will have more bacteria growth, the one + ampicillin or the one – ampicillin. Explain your reasoning.

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Uses of Biotechnology in Plants

Make Transgenic Plants toDelay ripening-ex. Flavor-Saver Tomatoes

Uses of Biotechnology in Plants

Improve nutritional content e.g. Golden Rice

Herbicide resistance Coffee

Uses of Biotechnology in Animals

Be a pharmaceutical factory produce a large amount of an otherwise

rare biological substance for medical useHormonesBlood clotting factorsVaccines

Q & A: Recombinant DNA1) Why is it useful to create bacteria that contain a new

gene?2) What is the purpose of restriction enzymes in this

process?3) What is the purpose of DNA ligase?4) Describe the 3 steps for making recombinant DNA.5) Human Growth Factor is a protein. Based on what

you’ve learned in this unit, what biochemical processes must take place before HGH will be produced by bacteria?