3.5 – Genetic Modification and Biotechnology
DNA profiling is a technique for identifying and identifying individuals based on their DNA
profiles. Satellite DNA — vast stretches of DNA made up of repeating elements called short tandem repeats
– appears in the non-coding portions of an individual's genome (STRs). Individuals will have varying
numbers of repeats at distinct satellite DNA loci, resulting in different DNA profiles. In criminal
investigations (forensics) and to settle paternity disputes, DNA profiling is commonly used. For both, the
technique is the same: A DNA sample is taken (for instance, from blood, sperm, or saliva) and amplified
using PCR. Satellite DNA (containing STR sequences) is sliced into fragments using specific restriction
enzymes. Due to the variable length of their short tandem repetitions, fragment length will differ between
people. Gel electrophoresis is used to separate the fragments, and the resulting profiles are compared. Gel
electrophoresis is a technique for separating and isolating proteins or DNA fragments based on their
mass/size. Samples are placed in a gel block, and an electric current is used to transport the samples through
the gel. Smaller samples are less hindered by the gel matrix and hence travel through the gel more quickly.
As they travel at different speeds, this causes samples of various sizes to separate. Using restriction
endonuclease, DNA can be sliced into fragments; fragment lengths will vary based on the DNA sample.Since
DNA is negatively charged due to the presence of a phosphate group on each nucleotide, fragments
separate.DNA samples are run over an agarose gel, and the size of the fragments is determined by comparing
them to industry standards. By integrating a corresponding radiolabelled hybridisation probe, transferring
the segregated sequences to a membrane, and then visualizing using autoradiography, specific sequences
can be identified. Proteins contain positive and negative regions and can be folded into a variety of forms
(affecting size) (no clear charge). To solve the nonlinear and impart a consistent negative charge to proteins,
they must first be treated with an anionic detergent (SDS). Protein samples are processed through a
polyacrylamide gel and their sizes are compared to industry standards. Target proteins are identified by
staining with specific monoclonal antibodies after separated proteins are transferred to a membrane (Western
blotting). The polymerase chain reaction (PCR) is a laboratory-based method of artificially duplicating
DNA. The PCR process is used to amplify vast amounts of a specific DNA sequence from a small sample
of DNA. Each reaction cycle doubles the amount of DNA produced; a 30-cycle PCR sequence produces
over 1 billion copies (230). In a thermal cycler, PCR uses temperature fluctuations to control the replication
process in three steps: Denaturation - The DNA sample is heated to 95°C for 1 minute to split it into two
single strands. Annealing - DNA primers bind to the target sequence's 3' ends. Elongation — A heat-tolerant
DNA polymerase (Taq) attaches to the primer and replicates the strand for 2 minutes at 72°C. Other
laboratory procedures are employed to separate and alter the sequences after huge quantities of DNA have
been generated. In a given organism, a gene specifies a certain trait by encoding for a specific polypeptide.
Because the genetic code is (nearly) universal, an organism can express a new trait by introducing the
relevant gene into its genome. Gene modification is the process of transferring genes from one species to
another, and the resulting new organism is known as a transgenic. In agriculture, genetically modified
organisms (GMOs) are utilized to increase crop yields and lower farming costs. The use of GM crops, on
the other hand, is a sensitive topic, as economic benefits must be balanced against environmental hazards.
By adding genes for specific proteins, vitamins, or vaccinations, GM crops can be used to improve human
nutritional standards. Furthermore, GM crops that are free of common natural allergies or poisons can be
created. However, some people may experience unexpected negative health effects if particular genes are
added or removed Currently, not all foods containing GM components are labeled, making it difficult for
consumers to make educated judgments about their use. Drought, cold, and salinity resistance genes can be
added to GM crops to help them grow in a wider range of situations. GM crops can be engineered to give
higher yields (crops can potentially grow larger and faster). GM crops may contain genes that delay the rate
of spoilage, resulting in GM foods with extended shelf lives. Genetically modified crops may be resistant to
particular viruses or produce insecticides (reducing need for the use of pesticides). Herbicide-resistant crops
can be utilized to make weed control more efficient (which compete with crops for soil nutrients). Farmers
will profit financially from increased yields, lower farming costs, and the capacity to farm more land. Patent
protection, on the other hand, permits biotech companies to limit the usage of seeds and force farmers to pay
inflated fees for them. The capacity to farm GM crops in a wider range of locations may lessen the
requirement for associated deforestation. In addition, the development of pest-resistant crops reduces the
amount of chemical insecticides emitted into the environment. However, by competing with native plant
life, GM crops have the potential to diminish biodiversity in a region. Furthermore, proteins or poisons
produced by GM crops may have a harmful impact on ecosystem creatures. Herbicide-resistant weeds and
grasses could emerge as a result of cross-pollination by GM crops. GM crops that contain pest poisons may
hasten the evolution of pest species that are resistant to them. Bt corn is a type of genetically modified maize
that contains an insecticide gene from the Bacillus thuringiensis bacteria. Certain types of larvae, particularly
the European corn borer, are killed by this insecticide, which would otherwise consume the crop. Concerns
have been expressed that the expansion of Bt corn may be affecting monarch butterfly survival rates. While
monarch butterfly larvae only eat milkweed, pollen from Bt maize may be carried by the wind to dust
neighbouring milkweeds. A preliminary study was undertaken in 1999 to look into the link between Bt corn
pollen exposure and monarch caterpillar survival rates: Milkweed leaves were coated with pollen from Bt
corn and given to monarch caterpillars (to simulate spread via wind). Caterpillars fed non-dusted leaves or
leaves dusted with non-GM pollen were compared to caterpillars given non-dusted leaves or leaves dusted
with non-GM pollen for growth and mortality.Caterpillars exposed to Bt pollen were shown to eat less, grow
more slowly, and die at a higher rate. Some scientists believe these findings are suspect because they do not
accurately reflect natural conditions: The amount of Bt pollen on the leaves was larger than what would be
found naturally (e.g. rain would diminish build up). The larvae's food was restricted (in the field, larva could
feasibly avoid eating pollen dusted leaves). A second study compared monarch butterfly survival rates in
relation to their proximity to Bt maize fields: When monarch larvae were placed on or near an actual Bt corn
field, there was no significant increase in mortality. As a result, it was established that Bt pollen offers no
major damage to monarch butterfly populations. Clones are collections of genetically identical organisms or
cells generated from a single parent cell. Asexually reproducing organisms make genetically identical clones.
There are also systems that allow sexually reproducing organisms to make clones (e.g. identical twins). The
generation of stem cells is required for cloning multicellular organisms (differentiated cells cannot form
other cell types). The procedure of somatic cell nuclear transfer can be used to generate stem cells from adult
tissue (SCNT). Somatic cell nuclear transfer is a technique for creating cloned embryos from differentiated
adult cells. Adult donors' somatic cells are extracted and grown (these cells are diploid and contain the entire
genome)
To make an enucleated egg cell, an unfertilized egg is retrieved from a female adult and its haploid nucleus
is removed. To generate a diploid egg cell (containing the donor's DNA), the enucleated egg cell is united
with the nucleus from the adult donor. The egg is then stimulated to divide and develop into an embryo with
the help of an electric current. The embryo is then inserted into a surrogate's uterus, where it will grow into
a genetic clone of the adult donor. Many animals can reproduce asexually and so have natural cloning
mechanisms. Asexual reproduction is used by all bacteria, the majority of fungus, and many protist species
to make genetic clones. While most plants reproduce sexually, they also have asexual reproduction
mechanisms (vegetative propagation). Asexual reproduction is possible in several animal species through a
variety of ways. Binary Fission is a type of nuclear fusion. The parent organism splits in half, resulting in
two genetically identical daughter organisms. Planaria (flatworms) use this method of cloning, but bacteria
and protists also use it (e.g. euglena, amoeba) Budding The parent organism's cells split out, resulting in a
tiny daughter organism that eventually separates from the parent. This cloning process is found in Hydra,
but it is also seen in many yeast species. Fragmentation - A detached component of the parent organism
gives rise to new creatures. Starfish and several species of annelid worms use this method of cloning. A stem
cutting is a section of a plant stem that can be removed and vegetatively propagated into a new independent
clone. Nodes are the points on a stem where a leaf, branch, or aerial root can grow; the space between nodes
is known as internodes. The lower nodes of stem cuttings are usually covered while the upper nodes are left
exposed when put in soil. Stem cutting is a typical technique for propagating plants quickly (including sugar
cane, grapes and roses).
Comparison of Reproductive and Therapeutic Cloning IMAGE FROM BIO-NINJA