In this lesson we

• examine the use of DNA technology in forensics, medicine, and agriculture (SB2f)

Terms and topics related to DNA Technology:– Biotechnology– cloning– DNA fingerprinting– genetic engineering– Human Genome Project– plasmid– recombinant DNA

What is biotechnology?• The use of living organisms to do

practical tasks.

• Early examples:– The use of microorganisms to make

cheese and wine– Selective breeding of livestock and crops– Production of antibiotics from

microorganisms– Production of monoclonal antibodies

Goal of biotechnology today:

• To find practical applications of DNA tecniques for the improvement of human health and food production– Making gene products using Genetic Engineering

– Uses in basic research – Medical uses. Diagnosis of disease – Making vaccines and other pharmaceutical

products – Forensic uses of DNA such as DNA fingerprinting – Agricultural uses such as making transgenic plants

DNA Technology

• What we can do with DNA and what it can do for us

DNA TECHNOLOGYDNA recombination or genetic engineering is the direct manipulation of genes for practical purposes

Recombinant DNA technology

• Refers to the set of techniques for combining genes from different sources in vitro( in a test tube) and transfering this DNA into a cell so it can be expressed.

• These techniques were first developed around 1975 and resulted in the appearance of the Biotechnology industry

The tools of recombinant DNA

• Plasmids

• Restriction enzymes

• Gel electrophoresis

• PCR ( polymerase chain reaction)

– Researchers can insert desired genes into plasmids, creating recombinant DNA

• And insert those plasmids into bacteria

Bacterium

Bacterialchromosome

Plasmid

1 Plasmidisolated

3 Gene insertedinto plasmid

2 DNAisolated

Cell containing geneof interest

DNAGene ofinterest

Recombinant DNA(plasmid)

4 Plasmid put intobacterial cell

Recombinantbacterium

5 Cell multiplies withgene of interest

Copies of proteinCopies of gene

Clone of cellsGene for pestresistanceinserted intoplants

Gene used to alter bacteriafor cleaning up toxic waste

Protein used to dissolve bloodclots in heart attack therapy

Protein used tomake snow format highertemperature

Figure 12.1

Genetic Engineering

• DNA from one species is inserted into another species.

• Ex. Human Insulin for diabetics is now made by bacteria cells!

•Therapeutic hormones– In 1982, humulin, human insulin produced

by bacteria• Became the first recombinant drug

approved by the Food and Drug Administration

Figure 12.7A

Restriction enzymes cut DNA at specific sequences in the DNA

Restriction enzymes = scissors

Recombinant DNA

Two pieces of DNA cut with the same restriction enzyme will be able to re-combine with each other.

Plants

• 1) Examples of transgenic plants with resistance to viruses…

potatoes, tomatoes, tobacco• 2) Examples of transgenic plants with resistance

to insects… corn, cotton • 3) resistance to herbicides • 4) slow down spoilage in tomatoes • 5) Extreme example – strawberries that are resistant to drought, salt, insects, viruses, cold and frost, and improved taste

Animals

• Bacteria now produce all of the following…– Human growth hormone (HGH) – – Human insulin (replaced cow and pig insulin for

human therapy )– Follicle-stimulating hormone – Factor VIII (replaced clotting factors taken from

human blood.)

Electrophoresis• Separation of charged molecules

in an electric field.• Nucleic acids have 1 charged

phosphate (- charge) per nucleotide. Separation based (mostly) on length: longer molecules move slower.

• Done in agarose gel matrix to stabilize.

• average run: 100 Volts across a 10 cm gel, run for 2 hours.

• Stain with ethidium bromide: bonds between DNA bases and fluoresces orange.

• Run alongside standards of known sizes to get lengths

Gel electrophoresis sorts DNA molecules by size

+ +

– –

Powersource

Gel

Mixture of DNAmolecules ofdifferent sizes

Longermolecules

Shortermolecules

Completed gel

Figure 12.10

– After digestion by restriction enzymes• The fragments are run through a gel

–

+

Longerfragments

Shorterfragments

x

w

y

z

y

1 2

Figure 12.11B

Crime

• Forensic science is the use of scientific knowledge in legal situations.

• The DNA profile of each individual is highly specific.

• The chances of two people having exactly the same DNA profile is 30,000 million to 1 (except for identical twins).

•DNA and Crime Scene Investigations– Many violent crimes go unsolved

• For lack of enough evidence

– If biological fluids are left at a crime scene• DNA can be isolated from them

– DNA fingerprinting is a set of laboratory procedures

• That determines with near certainty whether two samples of DNA are from the same individual

• That has provided a powerful tool for crime scene investigators

Investigator at oneof the crime scenes(above), Narborough,England (left)

DNA Fingerprinting1st-The DNA molecule is cut with restriction enzymes

2nd- we have to separate the fragments

This is done by a technique called gel electrophoresis

The DNA is placed on a tray filled with gel through which an electric current runs causing the fragments to move through the gel. The segments separate by how far they move in the gel according to size.

The DNA will form bands corresponding to the bases (and no two people have the same sequence of bases) in the gel which are unique for each individual. This is DNA fingerprinting

Example

• A violent murder occurred.

• The forensics team retrieved a blood sample from the crime scene.

• They prepared DNA profiles of the blood sample, the victim and a suspect as follows:

Does suspect DNA bands match DNA from the crime scene?

Suspects Profile

Blood sample from crime scene

Victims profile

– DNA fingerprinting can help solve crimes

Defendant’sblood

Blood fromdefendant’s clothes Victim’s

blood

Figure 12.12A Figure 12.12B

DNA Fingerprinting

Gel Electrophoresis separates pieces of

DNA based on size (after being cut up with

restriction enzymes)Different people will

have different banding patterns.

Related individuals will have similar patterns.

Solving Medical Problems

DNA profiles can be used to determine whether a particular person is the parent of a child.

A childs paternity (father) and maternity(mother) can be determined.

This information can be used in• Paternity suits• Inheritance cases• Immigration cases

Example: A Paternity Test

• By comparing the DNA profile of a

mother and her child it is possible to

identify DNA fragments in the child

which are absent from the mother and

must therefore have been inherited

from the biological father.

Does this man’s DNA match the DNA of the child?

Mother Child Man

29

Cloning organismsCloning organisms

• A body cellbody cell from one organism and an egg cellegg cell from another are fused

• The resulting cell divides divides like a normal embryolike a normal embryo

30

CloningCloning• CloneClone- a

member of a group of genetically identical cells

• May be produced by asexual asexual reproductionreproduction (mitosis)

What is cloning?

• Reproductive cloning– Making an exact copy of a

pre-existing or currently existing organism.

• Therapeutic cloning– Embryos are grown and

stem cells from the embryo are harvested to be used in research to develop treatment for cancer, Alzheimer’s, etc.

– Very controversial b/c it kills the embryo in the process.

Human Genome Project

• Started 1988, finished 2001, the entire sequence of bases in human DNA is now known.

• This multi-national effort has led to increased knowledge of …– Human genetic diseases– Gene therapies– Evolutionary relationships– Cellular functions– Cancer genes

Gene Libraries

• Human genes (and other genes of interest) can be stored inside bacteria cells and viruses which can be saved and grown for use in research.

• This may also preserve the genes of endangered or extinct species.

Review and Summarize

• -use of DNA technology in forensics, medicine, and agriculture (SB2f)

Terms and topics related to DNA Technology:– biotechnology– Cloning– DNA fingerprinting– genetic engineering– Human Genome Project– plasmid– recombinant DNA