12 GENETIC ENGINEERING AND THE FUTURE Flashcards
known to be the best known application of molecular genetics
Genetic engineering
Genetic engineering is made possible by?
- A method of **breaking and joining DNA **molecules derived from different source
forming the recombinant DNA. - A suitable gene carrier that can replicate both itself and foreign DNA segment attached
to it; - Means of introducing the composite DNA molecule or chimera into a functional host
cell; and - A method of selecting from a large population of cells clone of recipient cell has acquired the molecular chimera.
6 Requirements for Recombinant DNA Technology
- Restriction Endonuclease and DNA Ligase
- DNA ligase
- Vehicles for cloning
- Functional host cell
- A method for selection of recipient cells and their multiplication
- Recipient cells
part of cell’s defense mechanisms against invading foreign genomes
Restriction Endonuclease and DNA Ligase
cut within certain well-defined based sequences in the invading DNA fragments
Restriction Endonuclease and DNA Ligase
shown a high degree of specificity - possible to direct the splitting of DNA of the
cloning vehicle
Restriction Endonuclease and DNA Ligase
Enzyme that makes possible the insertion of selected piece of DNA
DNA ligase
Commonly used vector/vehicles for molecular cloning
a. Plasmids
b. Bacteriophages
c. Agrobacterium tumefaciens
The cloned DNA is introduced into the host cell (using ____).
Functional host cell
E. coli
Other host cells include:
Yeast
other bacteria
Drosophila
higher plants & animals
cells that show the characteristics of the introduced gene
Recipient cells
refers to the joining together of DNA molecules from
two different species that are inserted into a host organism to produce new genetic
combinations that are of value to science, medicine, agriculture, and industry
Recombinant DNA technology
e general name for a piece of DNA
that has been created by the combination of at least two strands
Recombinant DNA (rDNA),
6 Steps of Genetic Recombination Technology
- Isolation of Genetic Material
- Restriction Enzyme Digestion
- Amplification Using PCR
- Ligation of DNA Molecules
- Insertion of Recombinant DNA into the Host
- Isolation of Recombinant Cells
Since DNA exists within the cell membrane along with other macromolecules such as RNA,
polysaccharides, proteins, and lipids, it must be separated and purified which involves enzymes such
as ____
lysozymes, cellulase, chitinase, ribonuclease, proteases etc.
Other macromolecules are removable with other enzymes or treatments. Ultimately, the addition of
____ causes the DNA to precipitate out as fine threads. This is then spooled out to give purified
DNA
ethanol
act as molecular scissors that cut DNA at specific locations. These reactions
are called ____
Restriction Enzyme Digestion
‘restriction enzyme digestions’.
They involve the ____ with the selected restriction enzyme, at conditions
optimal for that specific enzyme
incubation of the purified DNA
The technique ____ reveals the progress of the restriction enzyme
digestion
This allows separating and cutting out the digested DNA fragments
‘Agarose Gel Electrophoresis’
On the application of current, the
negatively charged DNA travels to the positive electrode and is separated out based on size.
method of making multiple copies of a DNA
sequence using the enzyme – DNA polymerase in vitro.
Polymerase Chain Reaction or PCR
It helps to amplify a single copy or a few copies of DNA into thousands to millions of
copies.
PCR
PCR reactions are run on ‘thermal cyclers’ using the following components:
a) Template – DNA to be amplified
b) Primers – small, chemically synthesized oligonucleotides that are complementary to a
region of the DNA.
c) Enzyme – DNA polymerase
d) Nucleotides – needed to extend the primers by the enzyme.
The cut fragments of DNA can be amplified using PCR and then ligated with the cut vector
The process of joining these two pieces together using the enzyme?
‘DNA ligase’ is
‘ligation
The resulting DNA molecule is a hybrid of two DNA molecules – the interest molecule and
the vector.
This intermixing of different DNA strands is called
____.
recombination
new hybrid DNA molecule is also called a recombinant DNA molecule and the
technology is referred to as the
recombinant DNA technology
the recombinant DNA is introduced into a recipient host cell mostly, a
bacterial cell
Transformation
Bacterial cells do not accept foreign DNA easily. Therefore, they are treated to make
them ‘competent’ to accept new DNA. The processes used may be ____ etc.
thermal shock,
Ca++ ion treatment,
electroporation
The transformation process generates a mixed population of ____
transformed and non-transformed host cells.
The selection process involves filtering the ____ only
transformed host cells
For isolation of recombinant cell from non-recombinant cell, ____ is employed.
marker gene of plasmid vector
Genetic engineering has the following applications:
a) enhanced the field of biotechnology
b) opened new avenues perceived to be impossible before
DNA sequencing together with computer software that align DNA pieces facilitated the
shift from gene mapping to DNA sequencing.
1995
Data on human genome sequencing is shown in the public
1996
Sequencing of 99% of the human genome accomplished
2000
Progress in microarray technology (for investigating gene function and interactions)
2000 to present
The technique of DNA fingerprinting involves the use of:
a) Restriction Fragment Length Polymorphisms (RFLPs)
b) Variable Number of Tandem Repeats (VNTRs) or Minisatellites
regions of DNA that vary
among individuals.
Restriction Fragment Length Polymorphisms (RFLPs)
When probe for large enough number of RFLPs are used, the degree of accuracy is ____: the likelihood of a match between two unrelated individuals
is less than one in 100 million
very high
method of
DNA fingerprinting consider tandemly repeated sequences
Variable Number of Tandem Repeats (VNTRs) or Minisatellites
The number and size of
VNTR differ in different individuals. Each VNTR size is given a probability score
which is based on its frequency within a ____
reference human population.
genes
that control heritable diseases such as:
a) Osteoarthritis
b) Duchenne muscular dystrophy
c) Huntington’s disease
d) breast cancer disposition gene (BRCA 1 and BRCA 2)
It is claimed that aging may be caused by the buildup mutation of mitochondrial DNA.
Scientists in Australia and Japan observed that ____ much faster than
____
mitochondrial DNA mutation
chromosomal DNA.
Researches on Animal Genes
- Super-ovulation Hormone
- Rat Pancreatic Peptide Gene
- Transgenic Fishes
- Transgenic Livestock and Poultry
A recombinant ____ produced by Integrated
Genetics, Inc. causes superovulation in cattle.
bovine follicle-stimulating hormone (bFSH)
The multiple egg each bovine releases can be artificially inseminated and the embryos
implanted into surrogate mother cow.
Injection of pancreatic peptide into a strain of obese rats ____ feeding
and absorption of fats
reduced
Transgenic salmon
Carries foreign gene for growth hormone
Alongside the growth hormone gene, the gene for ____ was inserted
Observe to grow faster than non-transgenics
beta galactosidase
To improve the characteristics of livestock and poultry, ____ are
transferred to the animals.
growth hormones
Transgenic pigs
30% more efficient in feed conversion
Reach the marketable size ____ than their non-transgenic counterpart
7 week earlier
Researches on Plant Genes
- Genetically Modified for resistance to insect pest, disease & herbicides
- Genetically modified for improved nutritional qualities
- Genetically modified for special products
- Genetically modified for enhanced quality traits
Insertion of ____ gene for crystals toxic to Lepidopteran insects into canola, corn, cotton, potato, soybeans and tomato provide season-long protection against
Lepidopteran insect pests.
Bacillus thuringiensis (Bt)
Insertion of Virus coat protein genes provides resistance against ____
papaya ring spot virus
and banana bunchy top virus.
Insertion of bar gene from ____ into tomato, potato soybean, corn and tobacco provide resistance against herbicides.
Streptomyces hygrospicus or bxn from Klebsiella
Fatty acid modification, eg. soybean, canola and sunflower oil with high levels of ____
oleic
acids
____ gene introduced into rice
Methionine-rich
Rice varieties with ____ and amino acid levels than non-transgenic rice
20% higher protein
Genes for enzyme that converts sucrose to fructans are inserted into ____. It
yields 40% of their dry weight as fructans.
sugar beets
Golden rice - genes from daffodil flower and bacterium Erwinia uredovora, the source
of ____ enzyme to enable rice to synthesize ____, precursor of
vitamin A
phytoene desaturase
carotene
Gene for ____ (iron storage protein) - from soybean, then inserted to rice. ____
accumulates in the endosperm, tripling the rice iron content.
ferritin
Transgenic potatoes producing vaccine against ____
E.coli enterotoxin, cholera toxin and
Norwalk virus.
Transgenic bananas and tomatoes producing vaccines against ____
hepatitis B
Transgenic tomatoes producing vaccines against _____
respiratory syncytial virus
(causes lower respiratory tract disease in infant and children)
Genetically modified for enhanced quality traits
- Delayed tomato ripening
- Improved flavor and sweetness
- Preventing browning of apples & potatoes
Delayed tomato ripening by inserting antisense gene for ____
polygalacturonase (enzymes for breaking down pectin)
tomatoes with antisense gene ripen on the vine to develop flavor and color before the softening process, thus prolonging its shelf life.
Improved flavor and sweetness by inserting genes for ____ into tomato and lettuce
moneliin
Preventing browning of apples & potatoes by insertion of antisense gene for ____
polyphenol oxidase
(enzyme that polymerizes phenolic compound to form dark pigment)
Genome mapping requires identification of ____
DNA markers
DNA fragments that can distinguished from one another due to their
differences in nucleotide sequences.
DNA markers
used to leach metals (Cu, uranium) from Fe ore
Used to process waste H2S gas produced by petroleum refineries
Iron-oxidizing bacteria
GE microorganism that can track other GE organisms in the environment
Microbes to track organisms
Bacteria genetically engineered to glow in the dark
Can be used as living biosensors for detection of virtually any class of toxic agents
Bacterial biosensors
insulin is for
and treatment for?
Hormone for glucose uptake
For diabetis
tissue plaminogen activator TPA is for
and treatment for?
Dissolves blood clot
For heart attack victims
factor VIII is for
and treatment for?
Blood clotting factor
For certain types of
hemophilia patients
renin inhibitor is for
and treatment for?
Lower blood pressure
For hypertension
production of medically important proteins in the mammary gland of livestock
Molecular pharming
host of tissue plasminogen activator
use for?
goat
Dissolves blood clot
host of lactoferrin
use for?
cattle
Used as iron supplement in
infant formula
host of Insulin-like growth factor 1
use for?
cattle
Treatment of diabetis
The Release of Genetically Engineered Organisms: Biosafety and Ecological Implications
- Potential ecological effects
- Regulatory policies
Most GE organisms are modified domesticated species living under controlled
conditions. They are ____ human support
unable to survive without
Undesirable outcomes that should not be allowed to happen.
Gene flow through hybridization with related crops or wild species.
Damage to non-target species.
Disruptive effect on biotic communities.
Potential ecological effects
____ must be the basis for planning for the development, testing and
risk assessment of the introduction of GE organisms into environment
Ecological principles
The introduction of GE is directed at ____ while guarding against
potential problems.
promoting effectiveness
