I B B I O LO GY
4.4 GENETIC ENGINEERING AND BIOTECHNOLOGY
ASSESSMENT STATEMENTS
4.4.1 Outline the use of polymerase chain reaction (PCR) to copy and amplify minute quantities of DNA.4.4.2 State that, in gel electrophoresis, fragments of DNA move in an electric field and are separated according to size.4.4.3 State that gel electrophoresis of DNA is used in DNA profiling.4.4.4 Describe the application of DNA profiling to determining paternity and also in forensic investigations.4.4.5 Analyze DNA profiles to draw conclusions about paternity or forensic investigations.4.4.6 Outline three outcomes of the sequencing of the complete human genome.4.4.7 State that, when genes are transferred between species, the amino acid sequence of polypeptides translated from them is unchanged because the genetic code is universal.4.4.8 Outline a basic technique used for gene transfer involving plasmids, a host cell (bacterium, yeast or other cell), restriction enzymes (endonucleases) and DNA ligase.4.4.9 State two examples of the current uses of genetically modified crops or animals.4.4.10 Discuss the potential benefits and possible harmful effects of one example of genetic modification.4.4.11 Define ‘clone.’4.4.12 Outline a technique for cloning using differentiated animal cells.4.413 Discuss the ethical issues of therapeutic cloning.
GENETIC TECHNIQUES
We will be exploring ways in which scientists are able to explore and manipulate DNA.
Copying DNA in the laboratory (Polymerase Chain Reaction or PCR).
Using DNA to reveal its owner’s identity (DNA Profiling).
Mapping DNA by finding where every A, T, C and G is (The Human Genome Project).
Cutting and pasting genes to make modified organisms (Gene Transfer).
Cloning cells and plants/animals.
POLYMERASE CHAIN REACTION (PCR)
PCR is a laboratory technique which takes a very small quantity of DNA and copies it to make millions of copies
of DNA in only a few hours.
PCR is useful when small quantities of DNA are found in a sample but larger amounts are needed for analysis/testing.
DNA from very small samples of semen, blood, tissue, or long-since dead/extinct organisms can be amplified using PCR. PCR is carried out at high temperatures using a DNA
polymerase enzyme found from Thermus aquaticus, a bacterium living in hot springs.
PCR ANIMATION(S)
GEL ELECTROPHORESIS
This laboratory technique is used to separate fragments of DNA, proteins or other charged molecules in order to identify their origin. 1. Enzymes are used to chop up the long strands of DNA into
varying sized fragments. 2. The DNA fragments are placed into small wells (holes) in the gel
that are aligned along one end. The sheet of gel will act as a molecular sieve.
3. The gel is exposed to an electric current – positive on one side and negative on the other.
The biggest, heaviest and least charged particles do not move easily through the gel matrix—so they stay close to the wells they
were place in. The smallest, least massive, and most charge particles pass through the gel and move to the opposite side with little difficulty. Intermediate particles are distributed in between.
ELECTROPHORESIS RESULTS
GEL ELECTROPHORESIS
GEL ELECTROPHORESIS ANIMATIONS
DNA PROFILING/FINGERPRINTING
The use of gel electrophoresis to match an unknown sample of DNA with a known sample of DNA.
DNA Profiling is Used For:1. Paternity suits (identifying the biological father).2. Forensic investigations3. Evidence for evolutionary relationships
How are DNA Profiles Analyzed?
See Next Slide
DNA PROFILING (ANIMATIONS/VIDEOS)
FURTHER EXPLANATION(S)
DNA PROFILING LABS/INTERACTIVES
PDF File
THE HUMAN GENOME PROJECT(1990 – 2003)
The Human Genome Project was an international cooperative venture that set out to sequence the
complete human genome. The HGP hoped to determine all of the bases (A, T, C, G) in human DNA.
• Scientists are now actively determining which sequences represent protein-coding, genes. • The human genome can be thought of as a map
which can be used to show the locus of any gene on any one of the 23 pairs of chromosomes.• Browse our Genome
IMPORTANCE OF THE HUMAN GENOME PROJECT
1. Approximately 98.5% of our DNA is shared with chimpanzees (our closest living relatives).
2. Your DNA is 99.9% identical to the person sitting next to you. 3. The human genome consists of approximately 20,000-25,000
protein-encoding, genes.4. To a certain degree, ancestry can be mapped by comparing
the DNA of different populations around the world.Also . . . The Human Genome Can be Used To:5. Find beneficial molecules produced naturally by healthy
people. 6. Find out which genes control the synthesis of desirable
molecules/proteins.7. Copy that gene and use it as instructions to synthesize the
molecule(s) in the laboratory.8. Distribute the beneficial molecule as a new medical treatment.
CHROMOSOME 1
VIDEOS/ANIMATIONS
GENE TRANSFER
The genetic engineering technique of taking a gene out of one organism (donor) and placing it into another
organism (host).We an do this because:• We now have the technical and scientific knowledge
to do so.• All living things are based on a universal genetic code
(DNA). The same codons code for the same amino acids.
Examples:1. Bt-corn 3. Florescent
Plants/Animals2. Cold-resistant tomatoes 4. Insulin producing E.
coli
HOW IT WORKS: CUTTING, COPYING AND PASTING GENES (THE BASICS)
Although the laboratory techniques are complex, the concepts themselves are not difficult.
Cutting & Pasting DNA:• The molecular ‘scissors’ used for cutting bases sequences of
DNA are enzymes (restriction enzymes or endonucleases). These enzymes (normally used by bacteria kill pathogens) can find and locate specific sequences of bases pairs along a DNA molecule. Some can locate target sequences of four base pairs, others six base pairs. If both the beginning and end of the gene are cut, the gene is released and can be removed from the donor organism. For pasting genes, the enzyme used is called DNA ligase. It recognizes the parts of the base sequences that are supposed to be clicked together, called the sticky ends, and attaches them.
HOW IT WORKS: CUTTING, COPYING AND PASTING GENES (THE BASICS)
Copying DNA (DNA Cloning):• This is more complex because a host cell is needed in addition to the cutting
and pasting enzymes described in the previous slide. • Although yeast cells can be used as host cells, the most popular host cell in
genetic engineering is the bacterium Escherichia coli.• Like other prokaryotes, most of the genetic information for E. coli is in the
bacterium’s single chromosome. Some DNA, however, is found in structures called plasmids.
• Plasmids are small circles of extra copies of DNA floating around inside the cell’s cytoplasm. To copy a gene, it must first be glued into a plasmid.
To do this, a plasmid is removed from the host cell and cut using a restriction endonuclease. The gene to be copied is inserted into the open plasmid (gene
splicing). DNA ligase is used to paste the gene into the plasmid. The plasmid is now called a recombinant plasmid and it can be used as a vector (tool for
introducing a new gene into an organism’s genetic makeup. In the final step, the vector is placed inside the host bacterium and the bacterium is allowed to grow/divide in a bioreactor. The host cell (modified E. coli) will now express the
inserted gene and express the protein (insulin example).
HOW IT WORKS
GENETICALLY MODIFIED ORGANISMS
GMOs are organisms that have had one or more artificial genetic changes using the techniques of genetic engineering such as
gene transfer or recombinant DNA. • The simplest kind of genetic modifications are the removal of
undesirable genes or the addition of more desirable genes.
GENETICALLY MODIFIED (TRANSGENIC) ORGANISMS
Transgenic Plants:• The first commercial example of a GM food was the Flavr Savr
tomato sold in 1994. It was genetically modified to delay the ripening and rotting process so that it would stay fresher longer (food miles).
• Another plant of major interest is Golden Rice. This genetically modified rice plant has been engineered to produce beta carotene (used to produce Vitamin A) in the rice grains.
• Another genetically modified tomato was designed to withstand high salinity in the soil.
Transgenic Animals:• Hemophilia is a blood condition in which the blood does not clot
properly because of a lack of protein called factor IX. • The least expensive way of producing factor IX is to use transgenic
sheep. The gene for factor IX production is associated with the genetic information for producing milk. Female sheep will produce milk containing factor IX.
TRANSGENIC PLANTS
BENEFITS AND RISKS OF GENETIC MODIFICATION
Maize crops are often seriously damaged by corn borer insects. A gene from a bacterium (Bacillus thuringiensis) has ben transferred to maize. The gene codes for a
bacterial protein called Bt toxin that kills corn borers feeding on the maize.Potential Benefits of Bt Maize:1. Less pest damage and therefore higher crop yields to help reduce food shortages.2. Less land needed for crop production, so some could become areas for wildlife
conservation.3. Less use of insecticide sprays, which are expensive and can be harmful to farm
workers, consumers and wildlife.
Possible Harmful Effects of Bt Maize:4. Humans or farm animals that can eat the genetically modified maize might be
harmed by bacterial DNA in it, or by the Bt toxin.5. Insects that are not pests could be killed. Maize pollen containing the toxin is
blown onto wild plants growing near the maize. Insects feeding on the wild plants, including Monarch butterfly caterpillars, are therefore affected even if they do not feed on the maize.
6. Populations of wild plants might be changed. Cross-pollination will spread the Bt gene into some wild plants but not others. These plants would then produce the Bt toxin and have an advantage over other wild plants that struggle for survival, resulting in a reduction in biodiversity.
IS GENETIC ENGINEERING A GOOD OR BAD THING?
Genetic engineering raises many profound social and ethical questions.Benefits, Promises and Hopes:GM crops may help farmers improve food production. Is hunger a problem
of food production or food distribution?GM crops can produce beneficial substances like Vitamin A.GM crops which produce their own pest control substances will be
beneficial to the environment because fewer chemical pesticides will be used.
Using GMOs to produce rare proteins for medications or vaccines would be, in the long run, cheaper and produce less pollution than producing in the lab.
Farmers can be more in control of what crops or livestock they produce (maybe). There is always some randomness in breeding; genetic modification may make the process less of a gamble. It is also much quicker than selective breeding.
The multinational companies who make GM plants claim they will enable farmers in developing nations to help reduce hunger by using pest-resistant crops or GM plants requiring less water.
GENETICALLY MODIFIED ORGANISMS
IS GENETIC ENGINEERING A GOOD OR BAD THING?
Harmful Effects, Dangers and Fears: No one knows the long-term effects of GMOs in the wild. Efforts to keep GM
plants under control in well-defined areas have failed and pollen from GM crops has escaped to neighboring fields. Genes from GM plants could be integrated into wild species, giving then an unnatural advantage over other species and an ability to take over the habitat.
There is a danger that genes could cross species. It has been proven possible in laboratories. No one knows the consequences.
Bt crops which produce toxins to kill insects could be harmful to humans or other non-target insects/animals. The Bt toxin is found throughout the plant.
There are risks for allergies with GM crops (tomatoes). Most countries do not require labeling on GM crops.
Critics are worried that a large portion of the human food supply will be the property of a small number of very powerful corporations (Monsanto).
Critics are also worried that these very powerful corporations can bankrupt local farmers.
High-tech solutions are not necessarily better than simpler solutions. Crop production could be increased by teaching farmers how to use water/natural pest controls more effectively.
A proliferation of GMOs may lead to a decrease in biodiversity.
CLONES AND CLONING
Cloning: Producing identical copies of genes, cells or organisms.
The products of cloning are called clones. A clone is a group of genetically identical organisms or a group of
genetically identical cells derived from a single parent.
Cloning is very useful if an organism has a desirable combination of characteristics and more organisms with the same characteristic are wanted—this is reproductive cloning. Sometimes cloning is used to produce skin or other tissues needed to treat patient—this is called therapeutic cloning.
CLONING USING A DIFFERENTIATED ANIMAL CELL
In 1996, a sheep by the name of Dolly was born. She was the first clonewhose genetic material did not originate from an egg cell.
DOLLY WAS PRODUCED VIA REPRODUCTIVE CLONING.
Steps Involved1. From the original donor sheep to be cloned, a somatic (body or
non-gamete cell) from the udder was collected and cultured. The nucleus was removed from a cultured cell.
2. An unfertilized egg was collected from another sheep and its nucleus was removed.
3. Using a zap of electric current, the egg cell and the nucleus from the cultured somatic cell were fused together.
4. The new cell developed in vitro in a similar way to a zygote and started to form an embryo.
5. The embryo was placed in the womb of a surrogate mother sheep.
6. The embryo was developed normally.7. Dolly was born, an presented to the world as a clone of the
original donor sheep.
CLONING DOLLY THE SHEEP
CLONING USING UNDIFFERENTIATED CELLS (THERAPEUTIC CLONING)
In many cases, scientists are not interested in making a whole organism but interested in simply making copies of cells. This is called therapeutic cloning.
The aim of therapeutic cloning is to develop cells which have not yet gone through the process of differentiation.
This process generally involves using embryonic stem cells and stem cell research.
ETHICAL ISSUES SURROUNDING THERAPEUTIC CLONING
Since, in most cases, therapeutic cloning begins with the production of human embryos, it raises
fundamental ethical questions.
Is it ethically acceptable to generate a new human embryo for the sole purpose of medical research?
In nature, embryos are created only for reproduction and many people believe that using them for experiments is unnatural and morally wrong.
THERAPEUTIC CLONING
The use of embryonic stem cells, however, has led to major breakthroughs in our understanding of human biology.
How Can Therapeutic Cloning Help?:1. Embryonic stem cells can be used for therapies that save
lives and reduce suffering. 2. Growing skin to repair a serious burn.3. Growing new heart muscle to repair an ailing heart.4. Growing new kidney tissue to rebuild a failing kidney.5. Facilitating new connections between neurons to repair
spinal cord/brain injuries.
The vast majority of scientists, researchers and medical professionals are against the idea of reproductive human
cloning.
THERAPEUTIC CLONING
STEM CELL REFRESHER
THERAPEUTIC CLONING IN HUMANS
Arguments for Therapeutic Cloning:1. Embryonic stem cells can be used for therapies that save lives
and reduce human suffering.2. Cells can be removed from embryos that have stopped
developing and would have died anyway.3. Cells are removed at a stage when embryos have no nerve
cells and cannot feel pain.
Arguments Against Therapeutic Cloning:4. Every human embryo is a potential human being, which should
be given a chance of developing.5. More embryos may be produced than are needed, so some
may have to be killed.6. There is a danger of embryonic stem cells developing into
tumor cells during the therapeutic process.
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