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
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