GENE TECHNOLOGY - EXTENDED LECTURE OUTLINE

weedheightswaistBiotechnology

Dec 11, 2012 (4 years and 6 months ago)

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10
GENE TECHNOLOGY

EXTENDED LECTURE OUT
LINE

Genetic Engineering
(p. 222)

10.1

A Scientific Revolution

(p. 222; Fig. 10.1)

A.

Transferring genes from one organism to another falls into the realm of genetic engineering.

B.

Genetic engineering is having a major impact o
n medicine and agriculture.

10.2

Restriction Enzymes

(p. 223; Fig. 10.2)

A.

The first step of genetic engineering is to cleave the DNA that the geneticist wishes to
transfer.

B.

This process involves the use of restriction enzymes that bind specific sequences of
nuc
leotides and split the DNA in that position.

C.

Since DNA is made up of complementary bases, both strands do not split at the same
position.

D.

Instead, “sticky” ends result because the DNA is cleaved at an angle.

E.

These “sticky” ends can then be joined with a
ny other complementary sequence using ligase,
a sealing enzyme.

F.

Since only the ends are involved, the combining of DNA from different sources (i.e., human
and mouse, or human and bacteria) is possible.

10.3

The Four Stages of a Genetic Engineering Experiment

(
p. 224; Figs. 10.3, 10.4, 10.5, 10.6)

A.

Transferring genes from one organism to the next involves four stages.

B.

Stage 1: Cleaving DNA

1.

Cleaving DNA makes use of various restriction enzymes, each cleaving at a different
nucleotide sequence.

C.

Stage 2: Producing
Recombinant DNA

1.

Recombining DNA often makes use of a bacterial plasmid, a small circular piece of
DNA separate from the normal bacterial DNA.

D.

Stage 3: Cloning

1.

Cloning involves getting thousands of bacterial colonies to grow, which together make
up a clone
library.

E.

Stage 4: Screening

1.

The screening part of genetic engineering is often the most time
-
intensive, and
investigators must first eliminate any clones that do not contain vectors.

2.

Secondly, investigators use a probe of ribosomal RNA to detect the presen
ce of source
genes.

10.4

Working With DNA

(p. 228; Figs. 10.7, 10.8, 10.9)

A.

PCR Amplification

1.

Alternative to using bacterial plasmids to produce clones of source genes, geneticists use
the polymerase chain reaction (PCR) to produce many copies of the source gene
.

2.

This procedure involves locating short sequences of nucleotides, called primers, on either
side of the desired gene.

3.

Heat is applied to a solution of DNA, the primers, nucleotides, and DNA polymerase,
which disrupts the hydrogen bonds of DNA and produc
es single strands.

4.

When cool, the primers are bound to their complementary sequences near the desired
gene.

5.

The enzyme, DNA polymerase, then begins at a primer and replicates the single
-
stranded
DNA.

6.

Many copies of the desired gene can be made in this ma
nner.

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B.

Formation of cDNA

1.

Prokaryotes do not have exons interspersed with introns the way eukaryotes do, but
instead have continuous information encoded in the DNA.

2.

Thus, bacteria do not have the means to remove introns from eukaryotic DNA or newly
transcr
ibed mRNA.

3.

What researchers must do rather than using the primary transcript of mRNA is to isolate
the processed mRNA from the cytoplasm that eukaryotic cells have already excised of
introns.

4.

Reverse transcriptase is then used to make DNA from the process
ed mRNA, which
produces a type of gene called copy DNA (cDNA) that is made up only of exons.

5.

This cDNA can then be used by bacteria to produce proteins.

C.

DNA Fingerprinting

1.

DNA can be analyzed for its unique restriction endonuclease patterns, yielding
info
rmation that can be used to identify suspects from crimes.

Advances in Medicine

(p. 230)

10.5

Genetic Engineering and Medicine

(p. 230; Fig. 10.10, 10.11; Tables 10.1)

A.

Making “Magic Bullets”

1.

Medical advances have also been made since the advent of genetic engin
eering.

2.

Bacteria now mass
-
produce human insulin, the hormone that is underproduced in
diabetics.

3.

Other products, such as anticoagulants to dissolve blood clots and factor VIII to promote
clotting, are now safely produced by bacteria, which eliminates the

possibility of
transferring diseases from a human donor.

B.

Piggyback Vaccines

1.

Vaccines have been used for years to trigger immunity to a wide variety of diseases.

2.

Vaccines can now be made more safely by inserting the gene for a pathogen's surface
protein
into the DNA of a harmless virus.

3.

Such piggyback vaccines are being developed for malaria and other diseases.

C.

Human Gene Therapy

1.

Gene therapy, inserting normal genes into people who have inherited defective genes, is
now possible with the advent of geneti
c engineering.

Transforming Agriculture

(p. 233)

10.6

Genetic Engineering of Farm Animals

(p. 232; Figs. 10.12)

A.

Yet another agricultural advance has been the mass production by bacteria of bovine
somatotropin, which when fed to dairy cows, greatly enhances

milk production.

B.

Similar growth hormones enhance the size of pigs and cattle.

10.7

Genetic Engineering of Crop Plants

(p. 233; Figs. 10.14, 10.15; Table 10.2)

A.

Pest Resistance

1.

Certain crops, like cotton, have been engineered to be resistant to insect pests,

which
means these crops will not require pesticides.

2.

Bacterial genes that produce enzymes toxic to certain plant pests have been inserted into
tomatoes and other crops so that when the insect bites into a plant, it is killed by the now
plant
-
produced enz
ymes.

B.

Herbicide Resistance

1.

Agriculture has benefited from the genetic engineering of herbicide
-
resistant crops.

2.

The active ingredient in Roundup, called glyphosate, is easily broken down in the
environment and is thus a comparatively safe herbicide.

3.

Maki
ng crops resistant to glyphosate means less tilling is needed, thus soils are saved
from erosion and less fuel and expense are needed to raise the crop.

C.

More Nutritious Crops

1.

Rice can be modified to contain more minerals, such as iron, and vitamins.

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2.

“Gold
en” rice has been genetically engineered to contain vitamin A, a vitamin that is
normally insufficient in diets worldwide.

D.

How Do We Measure the Potential Risks of Genetically Modified Crops?

1.

Consumers worry that eating genetically
-
modified food might be d
angerous or that GM
crops are harmful to the environment.

2.

Other than allergic reactions to modified proteins, dangers to the consumer appear to be
slight.

3.

Whether GM products are potentially harmful to the environment is not yet clear.

a.

Wilmut removed mamma
ry cells from the udder of a six
-
year
-
old sheep and
combined them surgically with egg cells, then applied electrical shock to kick
-
start
the cell cycle.

b.

This cloning was the first successful cloning of an adult mammal.

E.

The Future of Cloning

1.

In 1997,

the same researchers that cloned the sheep cloned one with human genes in
every body cell.

2.

Transgenic cloning can be expected to have a major impact on medicine and agriculture.

3.

Cloning humans is an idea that is surrounded with controversy.


LEARNING OBJE
CTIVES



Recognize that the ability to manipulate genes and move them from one organism to another has led to
great advances in medicine and agriculture.



Explain how restriction enzymes are used.



List the four steps of a genetic engineering experiment.



Expla
in what clone libraries are and how cDNA probes can be used to screen for the presence of a
desired gene.



Discuss the uses and accuracy of DNA fingerprinting.



Give examples of how inserting human genes into bacteria has produced many medical advances.



Desc
ribe piggyback vaccines.



List ways in which agriculture has or will be transformed by genetic engineering.


KEY TERMS



genetic engineering

(p. 222) moving genes from one organism to another



restriction enzyme

(p. 223 ) Excising pieces of DNA is carried out
using restriction enzymes.



plasmid

(p. 225) A plasmid is a tiny ring of DNA in a bacterial cell that can replicate on its own.



clone library

(p. 225) All of the clones of the original DNA collectively are called the clone library.



probe

(p. 227) A probe i
s a complementary sequence of nucleotides that is used to locate the gene of
interest.



PCR

(p. 228) polymerase chain reaction



cDNA

(p. 228) copy DNA



factor VIII

(p. 230) One of the factors that leads to proper blood coagulation during a wound that is
missi
ng in one form of hemophilia.



piggyback vaccine

(p. 231) Making a piggyback vaccine involves inserting a gene for the microbe’s
surface protein into a harmless virus.


LECTURE SUGGESTIONS AND ENRICHMENT TIPS

1.

DNA Fingerprinting.

DNA fingerprinting employs
small bits of semen, hair, nails, skin, or blood taken
from a crime scene or from a victim of a crime. The DNA is isolated from these samples, and a
restriction enzyme is employed to cut repetitive portions of DNA into portions of varying length. How
long
the restriction fragments happen to be is unique to each person. The fragments are then sorted
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according to length using gel electrophoresis

small fragments migrate further. A membrane is used to
blot the electrophoresis gel and pick up a print of the gel.

Next, radioactively labeled DNA probes are
used to mark certain sequences of nucleotides within the suspects' DNA. The membrane is then used to
expose X
-
ray film, and a DNA fingerprint, with a unique series of banding patterns, is the result. No
two peopl
e have the same banding patterns, with the exception of identical twins. The probability that
two people chosen at random have the same banding pattern is on the order of one in a billion. DNA
fingerprinting is a very precise method of identification.

2.

How
Will Genetic Engineering Impact Traditional Agriculture?

If genetic engineering can produce
plants with complete proteins so humans have a diminished need to eat meat, what impact will this
have on traditional agriculture

for instance, on beef and pork pro
ducers? Will people still want to eat
beef, pork, chicken, and fish, or will they be willing to restrict their diets to plant nutrition only? Ask
students for their opinions on this matter. It is possible that animal protein will eventually become less
imp
ortant as plant protein is improved. Are there, however, other nutrients supplied by animal flesh
that one cannot get from plant sources in sufficient quantity?


CHANGES TO THE NEW EDITION

Refer to the Johnson instructor web site at
http://www.mhhe.com/biosci/genbio/tlw4

for a complete list of
changes to this edition.


CRITICAL THINKING QUESTIONS

1.

Explain why processed mRNA is used for the manufacture of a gene to be inserted into a bacterium.

2.

How does

cleaving DNA by using restriction enzymes facilitate the attachment of the cleaving DNA
into the plasmid?

3.

If it is possible to genetically engineer crops to fix their own nitrogen, comment on potential effects this
accomplishment will have on the world fo
od supply.


FILMS/MEDIA SUGGESTIONS

(Telephone and fax numbers and/or web sites for the sources of the following materials are listed in the
Appendix.)


Map of Life: Science, Society, and the Human Genome Project.

James Watson discusses this
fascinating, w
orldwide interaction to gather information about the human genome. Many of the people
interviewed also discuss bioethics and legal issues. 1992. 46 minutes.

CLEARVUE/eav,

WW5VH 1264


Genome: Solving the Code of Life.

How scientists find out about an organi
sm's genome is discussed,
along with the Human Genome Project. 30 minutes.

Carolina Biological Supply,

CE
-
53
-
6501


Animal Farm
. Transplanting animal organs into humans, with the help of genetic engineering, is the
subject of this video. Controversies surr
ounding this practice are discussed. 39 minutes.

Films for the Humanities and Sciences
, BVL7134


The Life Revolution
. This 12
-
part series of videos examines the nature of genetic science which may end
up transforming the way we live. Individual videos incl
ude:
Cutting and Splicing DNA; Evolution: Man
Takes a Hand; The Human Genome; DNA Techniques; Designer Plants; Depleting the Gene Bank; Sowing

the Seeds of Disaster; Growing Synthetics; Cell Wars; Recombinant Technology; Superanimals, Super
Humans?;
and

W
hither Biogenetics.

26 minutes each.

Films for the Humanities and Sciences
, BVL2430