Chapter 10

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14 Δεκ 2012 (πριν από 4 χρόνια και 6 μήνες)

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Biotechnology

Chapter 10

Golden Rice


Rice plants with added genes make and store
beta carotene



Video: Golden rice or Frankenfood?

GMOs and Transgenic Organisms




Transgenic


An organism that has been genetically modified
with genes from a different species



Genetically modified organisms

(GMOs)


Organism whose genome has been modified by
genetic engineering


10.2 Finding Needles in Haystacks



Gene research was limited until enzymes
produced by bacteria to cut viral DNA were
discovered



Restriction enzyme


Enzyme that cuts DNA at specific base sequences


Used in DNA cloning to cut DNA into pieces that
are inserted into cloning vectors

DNA Cloning


DNA cloning mass
-
produces DNA fragments for
research



DNA cloning


Set of procedures that uses living cells to make
many identical copies of a DNA fragment



Clone


A genetically identical copy of DNA, a cell, or an
organism

Cloning Vectors



Cloning vector


A DNA molecule that can accept foreign DNA,
resulting in a hybrid molecule that can be
transferred to a host cell, and get replicated in it



Plasmid


A small, circular DNA molecule in bacteria,
replicated independently of the chromosomes


A cloning vector


Recombinant DNA




Recombinant DNA molecules are introduced
into host cells such as bacteria, which copy the
DNA as they divide



Recombinant DNA


Contains genetic material from more than one
organism


Making Recombinant DNA

1. A restriction enzyme recognizes specific base
sequences in DNA from two different sources


2. Restriction enzymes cut DNA into fragments
with single
-
stranded tails (“sticky ends”)


3. DNA fragments from different sources are mixed
together; matching sticky ends base
-
pair


4. DNA ligase joins fragments, forming
recombinant DNA

Fig. 10
-
2, p. 181

restriction
enzyme (cut)

DNA ligase
(paste)

1

A restriction enzyme
recognizes a specific base
sequence in DNA (green
boxes) from two sources.

2

The enzyme cuts DNA
from both sources into
fragments that have
sticky ends.

3

The DNA fragments
from the two sources
are mixed together.

The matching sticky
ends base
-
pair with
each other.

4

DNA ligase joins

the fragments of
DNA where they
overlap. Molecules of
recombinant DNA

are the result.

mix

Making Recombinant DNA

Fig. 10
-
3, p. 181

Bam

Hl

Pst

l

Sph

l

Kpn

l

Eco

RI

Sal

l

Acc

l

Xho

l

Xba

l

Sac

l

Bst

XI

Not

l

Cloning Vector 3.85 kb

Plasmid

The use of mRNA for the
Identification of DNA


mRNA sequence comes from specific
regions of DNA (Genes)


mRNA sequence is used to make proteins
and defines the physical/behavioral
characteristics of the organism


Therefore we use mRNA to identify active
regions of DNA


Use mRNA sequence and base pairing
rules to identify DNA original sequence

cDNA Cloning


RNA cannot be cloned directly; reverse
transcriptase is used to copy single
-
stranded
RNA into cDNA for cloning



Reverse transcriptase


Viral enzyme that uses mRNA as a template to
make a strand of DNA



cDNA


DNA synthesized from an RNA template by the
enzyme reverse transcriptase

Making cDNA

Fig. 10
-
4, p. 182

mRNA

A The enzyme reverse
transcriptase transcribes

mRNA into DNA.

mRNA

cDNA

B DNA polymerase replicates
the DNA strand.

cDNA

cDNA

Eco

RI recognition site

C The result is a double
-
stranded molecule
of DNA that can be cut and pasted into a
cloning vector.

Stepped Art

Libraries



A library is a collection of cells that host different
fragments of DNA, often representing an
organism’s entire genome



Researchers make DNA libraries to isolate one
gene from the many other genes in a genome



Genome


An organism’s complete set of genetic material

Nucleic Acid Hybridization


Probes are used to identify one clone that hosts
a DNA fragment of interest among many other
clones in a DNA library



Probe


Short fragment of DNA labeled with a tracer


Hybridizes with a specific nucleotide sequence



Nucleic acid hybridization


Base
-
pairing between DNA or RNA from different
sources


PCR


PCR quickly mass
-
produces copies of a
particular DNA fragment for study



Polymerase chain reaction (PCR)


Uses primers and heat
-
resistant DNA polymerase
to rapidly generate many copies of a DNA
fragment



Primer


Short, single
-
strand of DNA designed to hybridize
with a DNA fragment

Steps in PCR

1. Starting material is mixed with DNA polymerase,
nucleotides and primers


2. Mixture is heated and cooled in cycles


At high temperature, DNA unwinds


At low temperature, primers base
-
pair with
template DNA


3.
Taq

polymerase synthesizes complementary
DNA strands on templates

2

When the mixture is heated, the
double
-
stranded DNA separates into
single strands. When it is cooled,
some of the primers base
-
pair with the
template DNA.

4

The mixture is heated again,

and the double
-
stranded DNA
separates into single strands. When
it is cooled, some of the primers
base
-
pair with the template DNA.

Fig. 10
-
5, p. 183

1

DNA template (
blue
) is mixed with
primers (
red
), nucleotides, and heat
-
tolerant
Taq

DNA polymerase.

Stepped Art

3

Taq

polymerase begins DNA
synthesis at the primers, and
complementary strands of DNA
form on the single
-
stranded
templates.

2

When the mixture is heated, the
double
-
stranded DNA separates into
single strands. When it is cooled,
some of the primers base
-
pair with the
template DNA.

5

Taq

polymerase begins DNA
synthesis at the primers, and
complementary strands of DNA
form on the single
-
stranded
templates.

4

The mixture is heated again,

and the double
-
stranded DNA
separates into single strands. When
it is cooled, some of the primers
base
-
pair with the template DNA.

Two Rounds of PCR

Animation: Polymerase chain reaction
(PCR)

Animation: Formation of recombinant
DNA

Animation: Use of a radioactive probe

Animation: Base
-
pairing of DNA
fragments

Animation: How to make cDNA

Animation: Restriction enzymes

Animation: F2 ratios interaction

10.3 Studying DNA




Short tandem repeats are multiple copies of a
short DNA sequence that follow one another
along a chromosome



The number and distribution of short tandem
repeats, unique in each individual, is revealed
by electrophoresis as a DNA fingerprint


DNA Fingerprinting



DNA fingerprinting is used in forensics, court
evidence, and other applications



DNA fingerprint


An individual’s unique array of short tandem
repeats



Electrophoresis


Used to separate DNA fragments by size

Fig. 10
-
6, p. 184

Evidence from
Crime Scene

Size Reference

Control DNA

Size Reference

Victim

Suspect 1

Suspect 2

Female Cells

Semen

Size Reference

Boyfriend

Control DNA

Control DNA

Size Reference

DNA Fingerprinting: A Forensic Case

The Human Genome Project




Automated DNA sequencing and PCR enabled
scientists to sequence the more than 3 billion
bases of the human genome



Sequencing


Method of determining the order of nucleotides in
DNA

Sequencing a Fragment of DNA



The order of colors is the order of DNA bases
(A, T, G, C)



Genomics




Analysis of the human genome sequence is
yielding new information about human genes
and how they work



Genomics


The study of genomes (structural genomics,
comparative genomics)


Some Sequenced Genomes

Animation: Automated DNA sequencing

Animation: DNA fingerprinting

Video: ABC News: DNA mystery: Human
chimeras

Video: ABC News: Family ties: Paternity
testing

3D Animation: Gene sequencing

10.4 Genetic Engineering


Recombinant DNA technology and genome
analysis are the basis of genetic engineering



Genetic engineering is the directed alteration of
an individual’s genome, resulting in a genetically
modified organism (GMO)



Genetic engineering


Process by which deliberate changes are
introduced into an individual’s genome

Genetically Modified Microorganisms



A transgenic organism carries a gene from a
different species



Transgenic organisms are used in research,
medicine, and industry



Transgenic bacteria and yeast produce
medically valuable proteins


Designer Plants




Transgenic crop plants help farmers produce
food more efficiently



Plants with modified or foreign genes are now
common in farm crops

Fig. 10
-
8, p. 187

1

An
A. tumefaciens
bacterium
has been engineered to contain a

Ti plasmid. The plasmid carries a
foreign gene.

2

The bacterium infects a plant
cell and transfers the Ti plasmid
into it. The plasmid DNA becomes
integrated into one of the cell’s
chromosomes.

3

The plant cell divides, and its
descendants form an embryo.

4
The embryo develops into

a transgenic plant.

5

The transgenic plant
expresses the foreign
gene. This tobacco plant is
expressing a gene

from a firefly.

Using the Ti plasmid

to Make a Transgenic Plant

Animation: Gene transfer using a Ti
plasmid

Genetically Modified Crops


Bt

gene confers insect resistance to corn



Biotech Barnyards



Transgenic animals produce human proteins



Animals that would be impossible to produce by
traditional breeding methods are being created
by genetic engineering



Transgenic animals are used in research,
medicine, and industry

Transgenic Animals

Knockout Cells and Organ Factories




Transgenic animals may one day provide a
source of organs and tissues for transplantation
into humans



Xenotransplantation


Transplant of an organ from one species to
another

Animation: Transferring genes into
plants

Video: ABC News: Cloned pooch

Video: ABC News: Mule clones

Video: ABC News: Glow
-
in
-
the
-
dark pigs

Video: ABC News: Cloned food approved

10.5 Genetically Modified Humans



Genes can be transferred into a person’s cells to
correct a genetic defect or treat a disease



However, the outcome of altering a person’s
genome remains unpredictable



Gene therapy


Transfer of a normal or modified gene into an
individual with the goal of treating a genetic
defect or disorder

Unpredictable Outcomes



There are more than 15,000 serious genetic
disorders


gene therapy is the only real cure



In some cases, gene therapy works


in other
cases it leads to death


Inserting a virus
-
injected gene into a
chromosome can disrupt normal function and
cause cancer


Severe allergic reaction to the viral vector can
cause death

One Successful Case of Gene Therapy


Rhys Evans, born with a severe immune
disorder (SCID
-
X1) received a normal gene and
no longer lives in isolation




Getting Perfect




Eugenics


Idea of deliberately improving the genetic
qualities of the human race



The potential benefits of genetically modifying
humans must be weighed against the potential
risks, including social implications

10.6
Impacts/Issues

Revisited




Golden rice with beta carotene was ready for
distribution in 2005 but is still not available for
human consumption


the biosafety experiments
required are too expensive for the public
humanitarian agency that developed it

Digging Into Data:

Enhanced Spatial
Learning in Mice With Autism Mutation