One cell with the recombinant plasmid

drawerbeamerBiotechnology

Dec 6, 2012 (5 years and 1 month ago)

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Biotechnology

Dolly and
surrogate Mom

Genetically modified rice.

Embryonic stem cells and
gene therapy

Biotechnology

Biotechnology, defined broadly, is the engineering of organisms for useful
purposes.

Often, biotechnology involves the creation of hybrid genes and their introduction
into organisms in which some or all of the gene is not normally present.

Fourteen month
-
old genetically engineered (“biotech”) salmon (left) and
standard salmon (right).

Biotechnology

We’ll examine:

Animal cloning

Gene cloning for
pharmaceutical production

The promise and perhaps
perils of embryonic stem cells

DNA fingerprinting

Genetically modified
foods and the American
-
European opinion divide.

Animal Cloning

Dolly and her surrogate mother.

Why Clone Animals?

To answer questions of basic biology

Five genetically identical cloned pigs.

For herd improvement.

To satisfy our desires (e.g. pet
cloning).

For pharmaceutical production.

Is Animal Cloning Ethical?

The first cloned horse and her surrogate
mother/genetic twin.

As with many important questions, the answer is beyond the scope of
science.

USU’s Contribution


A Cloned Mule and
the First Cloned
Equine

The Biotechnology
of Reproductive
Cloning

Even under the best of
circumstances, the
current technology of
cloning is very
inefficient.

Cloning provides the
most direct
demonstration that all
cells of an individual
share a common
genetic blueprint.

Saved by Cloning?

Some are firm believers while
many view these approaches to be
more of a stunt.

Note the use of a closely related
species, a domestic goat, as egg
donor and surrogate mother.

(Science (2002) 295:1443)

Carbon Copy


the First Cloned Pet

Significantly, Carbon Copy is not a phenotypic carbon copy of the
animal she was cloned from.

The Next Step?

Highly unlikely.

Attempts at human cloning are viewed very unfavorably in the scientific community.

Recombinant DNA, Gene Cloning, and Pharmaceutical Production

DNA can be cut at specific sequences using restriction enzymes.

This creates DNA fragments useful for gene cloning.

These are mature and widely utilized biotechnologies.

Restriction Enzymes are Enzymes That Cut DNA Only at
Particular Sequences

The enzyme EcoRI cutting DNA at its recognition sequence

Different restriction enzymes have different recognition sequences.

This makes it possible to create a wide variety of different gene fragments.

Restriction
enzyme
animation

DNAs Cut by a Restriction Enzyme Can be Joined Together
in New Ways

These are recombinant DNAs and they often are made of DNAs from
different organisms.

Plasmids are Used to Replicate a Recombinant DNA

Plasmids are small circles of DNA found in bacteria.

Plasmids replicate independently of the bacterial chromosome.

Replication often produces 50
-
100 copies of a recombinant
plasmid in each cell.

Pieces of foreign DNA can be added within a plasmid to create a
recombinant plasmid.

Harnessing the Power of Recombinant DNA Technology


Human Insulin Production by Bacteria

Human Insulin Production by Bacteria

6) join the plasmid and human fragment

and cut with a restriction enzyme

Human Insulin Production by Bacteria

Mix the recombinant plasmid with
bacteria.

Screening bacterial cells to learn which contain the human insulin gene is the
hard part.

Route to the Production by Bacteria of Human Insulin

A fermentor used to grow
recombinant bacteria.

This is the step when gene cloning takes place.

The single recombinant plasmid replicates within a cell.

Then the single cell with many recombinant plasmids produces trillions of
like cells with recombinant plasmid


and the human insulin gene.

One cell with the
recombinant plasmid

Route to the Production by Bacteria of Human Insulin

The final steps are to collect the bacteria, break open the cells, and purify the
insulin protein expressed from the recombinant human insulin gene.

Route to the
Production by
Bacteria of
Human Insulin

Overview of gene
cloning.

Cloning
animation

Pharming

These goats contain the human gene for a clot
-
dissolving
protein that is produced in their milk.

Pharming is the production of pharmaceuticals in animals engineered to
contain a foreign, drug
-
producing gene.

The Promise and Possible Perils of Stem Cells

The Stem Cell Concept

A stem cell is an undifferentiated,
dividing cell that gives rise to a
daughter cell like itself and a
daughter cell that becomes a
specialized cell type.

Stem Cells are Found in the Adult, but the Most Promising Types of
Stem Cells for Therapy are Embryonic Stem Cells

The Inner Cell Mass is the Source of Embryonic Stem Cells

The embryo is destroyed by separating it into individual cells for the collection
of ICM cells.

Some Thorny Ethical
Questions

Is it ethical to harvest
embryonic stem cells from
the “extra” embryos created
during
in vitro

fertilization?

Are these masses of cells a
human?

Additional Potential Dilemmas


Therapeutic Cloning to Obtain
Matched Embryonic Stem Cells

Cells from any source other than you or an identical twin present the
problem of rejection.

If so, how can matched embryonic stem cells be obtained?

A cloned embryo of a person can be made, and embryonic stem cells
harvested from these clones.

Cultured mouse embryonic stem cells.

Therapeutic Cloning

Is there any ethical difference between therapeutic and reproductive cloning?

DNA, the Law, and Many Other Applications



The Technology of DNA Fingerprinting

A DNA fingerprint used in a murder case.

What are we looking at? How was it produced?

The defendant stated that the blood on his clothing was his.

DNA Fingerprinting Basics

Different individuals carry different alleles.

Most alleles useful for DNA fingerprinting differ on the basis of the
number of repetitive DNA sequences they contain.

DNA Fingerprinting Basics

If DNA is cut with a restriction
enzyme that recognizes sites on either
side of the region that varies, DNA
fragments of different sizes will be
produced.

A DNA fingerprint is made by analyzing
the sizes of DNA fragments produced
from a number of different sites in the
genome that vary in length.

The more common the length variation at
a particular site and the greater the
number the sites analyzed, the more
informative the fingerprint.

A Site With Three Alleles Useful for DNA Fingerprinting

DNA fragments of different size will be produced by a restriction
enzyme that cuts at the points shown by the arrows.

The DNA Fragments Are Separated on the Basis of Size

The technique is gel electrophoresis.

The pattern of DNA bands is compared between each sample loaded on
the gel.

Gel electrophoresis animation

Possible Patterns for a Single “Gene” With Three Alleles

In a standard
DNA
fingerprint,
about a dozen
sites are
analyzed, with
each site
having many
possible
alleles.

A DNA Fingerprint

When many genes are
analyzed, each with many
different alleles, the
chance that two patterns
match by coincidence is
vanishingly small.

DNA detective
animation

HGP fingerprinting
page

DNA and the Law

SLT 3/8/05

Some applications of DNA
fingerprinting in the justice system.

Genetically Modified Foods

Many of our crops in the US are genetically modified.

Should they be?

GM Crops are Here Today

Source: Pew Initiative on Food and Biotechnology, August 2004.

Methods for Plant Genetic Engineering are Well
-
Developed and
Similar to Those for Animals

Golden Rice is Modified to be Provide a Dietary Source of Vitamin A

Worldwide, 7% of children suffer vitamin A deficiency, many of them living in
regions in which rice is a staple of the diet.

Golden rice (yellow)
with standard rice
(white).

Genetically Modified Crops

Genetically Modified Cotton

(contains a bacterial gene for
pest resistance)

Standard Cotton

GMOs, Especially Outside the US, Are a Divisive Issue

Protesters at the 2000 Montreal World
Trade Summit

European sentiment

Current Concerns by Scientists Focus on Environmental, Not
Health, Effects of GM Crops

The jury’s still out on the magnitude of GM crop’s ecological impact, but the
question is debated seriously.

Current Concerns
by Scientists
Focus on
Environmental,
Not Health,
Effects of GM
Crops