Techniques in Biotechnology: Ma. Joy O. Jocosol Eastern Samar State University

Chapter 2
Techniques in Biotechnology
MA. JOY O. JOCOSOL
EASTERN SAMAR STATE UNIVERSITY
• Tissue Culture
• DNA Extraction
• DNA Cloning
TOPICS • Gene Coding
• Gene Gun and Alternative
Techniques
• Backcross Breeding
• Cloning
• DNA Sequencing
2021 Techniques in Biotechnology 2

INTRODUCTION

INTRODUCTION

INTRODUCTION

INTRODUCTION

OBJECTIVES
At the end of this topic, we are expected to:

1. Understand the basic techniques in
biotechnology;
2. Enumerate the steps of each technique;
3. Appreciate the importance of these
techniques.
1. TISSUE CULTURE
Tissue culture, a method of

biotechnology, in which fragments of
tissue from an animal or plant are
transferred to an artificial environment
in which they can continue to survive
and function. The cultured tissue may
consist of a single cell, a population of
cells, or a whole or part of an organ.
Cells in culture may multiply; change
size, form, or function; exhibit
specialized activity (muscle cells, for
example, may contract); or interact
with other cells.
1. TISSUE CULTURE
Plant tissue culture is an essential component of plant biotechnology.
Apart from mass multiplication of elites, it also provides the means to
multiply and regenerate novel plants from genetically engineered cells.
The promising plant thus produced may be readily cloned in cultures
under aseptic conditions. Tissue Culture is widely used in:
Obtaining disease free plants.
Rapid propagation of plants those are difficult to propagate.
Somatic hybridization.
Genetics improvement of commercial plants.
Obtaining androgenic and gynogenic haploid plants for breeding
programs.
1. TISSUE CULTURE
Tissue Culture is becoming as an alternative means to vegetative
propagation of plants. In vitro growing plants are usually free
from bacterial and fungal diseases. Virus eradication and
maintenance of plants in virus free stage can also be rapidly
achieved in cultures. Three main methods generally used in
tissue culture are –
Micro propagation through the enhanced multiplication of
axillary bud.
 Organogenesis.
 Somatic embryogenesis.
1. TISSUE CULTURE
Importance of tissue culture
 In a relatively short time and space a large number of plantlets can be produced starting from the single
explant.
 Taking an explant does not usually destroy the mother plant, so rare and endangered plants can be cloned
safely.
 It is easy to select desirable traits directly from the culture setup (in vitro) thereby decreasing the amount
of space required, for field trials.
 Once established, a plant tissue culture line can give a continuous supply of young plants throughout the
year.
 The time required is much shortened, no need to wait for the whole life cycle of seed development. For
species that have long generation time, low level of seed production, or seeds that readily do not germinate,
rapid propagation is possible.
 In vitro growing plants usually free, from the bacterial and fungal diseases. Virus eradication and
maintenance of plants in virus free state. This facilitates movement of plant across international boundaries.
 Plant tissue banks can be frozen and then regenerated through tissue culture. It preserves the pollen and
cell collections from which plants may be propagated.

1. TISSUE CULTURE
Types of tissue culture
Callus culture: Callus culture may be defined as production and maintenance of an unorganized mass of proliferative cell from
isolated plant cell, tissue or organ by growing them on artificial nutrient medium in glass vials under controlled aseptic conditions.
Organ culture: That may allow differentiation and preservation of the architecture. The organ culture refers to the in vitro culture
and maintenance of an excised organ primordial or whole or part of an organ in way and function.
Single cell culture: Single cell culture is a method of growing isolated single cell aseptically on nutrient medium under controlled
condition.
Suspension culture: Suspension culture is a type of culture in which single cell or small aggregates of cell multiply while
suspended in agitated liquid medium. Suspension cultures are used in induction of somatic embryos and shoots, production of
secondary metabolites, in vitro mutagenesis, selection of mutants and genetic transformation studies.
Embryo culture: Embryo culture may be defined as aseptic isolation of embryo (of different developmental stages) from the bulk
of maternal tissue of mature seed or capsule and in vitro culture under aseptic and controlled physical condition in glass vials
containing nutrient semisolid or liquid medium to grow directly into plantlet.
Anther culture: Androgenesis is the in vitro development of haploid plants originating from potent pollen grains through a series
of cell division and differentiation.
Pollen culture: Pollen culture is the in vitro technique by which the pollengrains (preferably at the microscope stages) are
squeezed from the intact anther and then cultured on nutrient medium where the microspores without producing male gametes.
Somatic Embryogenesis: Somatic embryogenesis is the process of a single or group of cells initiating the development pathway
that leads to reproducible regeneration of non zygotic embryos capable of germinating to form complete plants.

1. TISSUE CULTURE
Protoplast Culture: It is the culture of isolated protoplasts which are naked plant cells surrounded by plasma membrane which is potentially
capable of cell wall regeneration, cell division, growth and plant regeneration on suitable medium under aseptic condition
Shoot tip and Meristem culture: The tips of shoots (which contain the shoot apical meristem) can be cultured in vitro producing clumps of
shoots from either axillary or adventitious buds. This method can, be used for clonal propagation.
Explant Culture: There are variety of forms of seed plants viz., trees, herbs, grasses, which exhibit the basic morphological units i.e. root,
stem and leaves. Parenchyma is the most versatile of all types of tissues. They are capable of division and growth.
Plant in vitro culture techniques

The promise of plant in vitro technologies in three major areas, namely micro propagation, somatic cell genetics and generation of transgenic
plant.
Micropropagation – Propagation in tissue culture (micropropagation) is, used to develop high-quality clonal plants (Smith 1990). The main
advantages are attributed to the potential of rapid, large scale propagation of new genotypes, the use of small amount of original germplasm.
Somatic cell genetics- Contribution of in vitro methods to plant breeding i.e. somatic cell genetics is most significant, mostly in terms of
haploid production and somatic hybridization.
Transgenic plants- Expression of mammalian genes or other plant gene is becoming routine for several plant species. One of the successful
approaches has been engineered for resistance against insects, virus and other pathogens as well as herbicide.
Biodiversity and strategies to preserve biodiversity- Biodiversity in common place is defining as species richness in given habitat.
Biodiversity exits as three major levels. Genetic biodiversity, species biodiversity and ecosystem biodiversity (Daniels, 1997)

2. DNA EXTRACTION
DNA Extraction is a
technique in
biotechnology which
involves isolation of
DNA in an organism.

2. DNA EXTRACTION
• Importance
The extraction of DNA is pivotal to biotechnology. It is
the starting point for numerous applications, ranging
from fundamental research to routine diagnostic and
therapeutic decision-making. Extraction and purification
are also essential to determining the unique
characteristics of DNA, including its size, shape and
function.
2. DNA EXTRACTION
• DNA was first isolated by the Swiss physician, Friedrich Miescher, in
1869 while working in the laboratory of the biochemist Felix Hoppe-
Seyler. This he did as part of a project to determine the chemical
composition of cells which he saw as the means to unravelling the
fundamental principles of the life of cells. Initially, he began this research
by using lymphocytes drawn from lymph nodes but was unable to get
sufficient quantities for analysis so switched to using leucocytes, white
blood cells, which he gathered from pus found on fresh surgical
bandages collected from a nearby surgical clinic. In the course of his
work on leucocytes he noticed the precipitation of a substance when
acid was added and that this dissolved following the addition of alkali.

2. DNA EXTRACTION
. Mierscher decided to call the new substance 'nuclein' by virtue of its presence in the
nuclei of the cell. Upon further analysis, Miescher noted that the chemical composition of
the substance differed from proteins and other known molecules. He speculated that it
played a central role in cells and was involved in the division of cells. Following this,
Miescher developed a method for isolating nuclein from salmon sperm. Many advances
have been made to the methods for extracting and purifying DNA since Miescher's time.
From the 1950s routine laboratory extraction of DNA became reliant on the use of
density gradient centrifuges. Until very recently most methods for extracting DNA
remained complex, labour-intensive and time-consuming. They also provided only small
quantities of DNA. Today there are many specialised extraction methods. These are
generally either solution-based or column-based. The extraction of DNA has become
much easier with the emergence of commercial kits and the automation of the process.
Such changes have both sped up production and increased the yield of DNA.

2. DNA EXTRACTION
Application
The ability to extract DNA is of primary importance to
studying the genetic causes of disease and for the
development of diagnostics and drugs. It is also
essential for carrying out forensic science,
sequencing genomes, detecting bacteria and viruses
in the environment and for determining paternity.

3. DNA CLONING
Cloning is the process

of moving a gene from
the chromosome it
occurs in naturally to an
autonomously
replicating vector..

3. DNA CLONING
In the cloning process, the DNA is removed from cells,

manipulations of the DNA are carried out in a test-
tube, and the DNA is subsequently put back into cells.
Because E. coli is so well characterized, it is usually
the cell of choice for manipulating DNA molecules.
Once the appropriate combination of vector and
cloned DNA or construct has been made in E. coli, the
construct can be put into other cell types
3. DNA CLONING
Deoxyribonucleic acid (DNA) cloning is a cornerstone of molecular biology. Its
power is such that without it a large proportion of modern biological science
would simply not be possible. A clone can be defined as an identical copy.
DNA cloning involves joining, or ‘ligating’, DNA with a ‘vector’ that enables the
resulting construct to be introduced into a cell, replicated and passed on to
daughter cells as that cell divides. The result is a population of cells all
containing clones of the original DNA. Thereafter that DNA is immortalized,
since cells can be grown to order and DNA extracted for study or further
manipulation whenever it is required. DNA cloning has been made possible by
the discovery of two types of protein, those that break or modify DNA such
that they have suitable termini for ligation and those that are capable of
ligating those molecules of DNA.
3. DNA CLONING
Isolating DNA from cells
Plasmid DNA isolation
The first step in cloning is to isolate a large amount of the vector and chromosomal DNAs. Isolation of
plasmid DNA will be examined first. In the general scheme, cells containing the plasmid are grown to a high
cell density, gently lysed, and the plasmid DNA is isolated and concentrated. When the cells are growing,
the antibiotic corresponding to the antibiotic resistance determinant on the plasmid is included in the growth
media. This ensures that the majority of cells contain plasmid DNA. Without the antibiotic selection, an
unstable plasmid (i.e. one without a par function) can be lost from the cell population in a few generations.
Chromosomal DNA isolation
To isolate chromosomal DNA, cells are lysed in much the same way as for plasmid DNA isolation. The cell
lysate is extracted with phenol or otherwise treated to remove all of the proteins. The chromosomal DNA is
precipitated as long threads. The chromosomal DNA is very fragile and breaks easily. For these reasons,
the chromosomal DNA is not usually purified using columns. Rather, the precipitated threads are collected
by centrifugation.

3. DNA CLONING
Cutting DNA molecules
Once DNA has been purified, it must be cut into pieces before the chromosomal DNA
and the plasmid DNA can be joined. The problem is to cut the DNA so that it will be
easy to join the cut ends of the chromosomal DNA to the cut ends of the plasmid
DNA. A group of enzymes, called restriction enzymes, are used for this purpose.
Restriction enzymes are isolated from different bacterial species.
Joining DNA molecules
Both plasmid and chromosomal DNA can be independently isolated from cells and
digested with restriction enzymes. If, however, DNA with double stranded ends is
simply transformed back into E. coli, E. coli will degrade it. The double-stranded ends
must be covalently attached. A version of this reaction is normally carried out in the
cell by an enzyme known as DNA ligase.

4. GENE CODING
Genetic code is the term we use for the

way that the four bases of DNA--the A,
C, G, and Ts--are strung together in a
way that the cellular machinery, the
ribosome, can read them and turn them
into a protein. In the genetic code, each
three nucleotides in a row count as a
triplet and code for a single amino acid.
So each sequence of three codes for an
amino acid. And proteins are made up of
sometimes hundreds of amino acids. So
the code that would make one protein
could have hundreds, sometimes even
thousands, of triplets contained in it.

4. GENE CODING
Life’s genetic code is written in the DNA molecule (aka deoxyribonucleic
acid). From the perspective of design, there is no human language that
can match the simplicity and elegance of DNA. But from the perspective
of implementation—how it is actually written and spoken in practice—
DNA is a linguist’s worst nightmare. DNA has four major functions: (1) it
contains the blueprint for making proteins and enzymes; (2) it plays a role
in regulating when the proteins and enzymes are made and when they
are not made; (3) it carries this information when cells divide; and (4) it
transmits this information from parental organisms to their offspring. In
this chapter, we will explore the structure of DNA, its language, and how
the blueprint becomes translated into a physical protein.

4. GENE CODING

4. GENE CODING

4. GENE CODING

4. GENE CODING
DNA replication
Complementary base pairing also assists in the faithful reproduction of the
DNA sequence, a process geneticists call DNA replication. When a cell
divides, both of the daughter cells must contain the same genetic
instructions. Consequently, DNA must be duplicated so that one copy ends
up in one cell and the other in the second cell. Not only does the replication
process have to be carried out, but it must be carried out with a high degree
of fidelity. Most cells in our bodies—neurons being a notable exception—are
constantly dying and being replenished with new cells. For example, the
average life span of some skin cells is on the order of one to two days, so the
skin that you and I had last month is not the same skin that we have today.

4. GENE CODING
DNA Packaging
To appreciate the way that DNA is packaged, we should first think of three types of
objects—a very, very long piece of twine, several million jelly donuts, and the Eiffel
tower. Taking the twine in one hand and a donut in the other, wrap the twine twice
around the donut in the way shown in panel. Leaving a few inches of twine free, grab
another donut and loop the twine around it twice. Repeat this process until you have
about six donuts with twine wrapped around them, giving a structure similar to that in
panel (b). Arrange these donuts in a circle on the ground right next to the mechanisms
that serve two purposes—helping to insure that DNA is copied accurately and
preventing DNA from becoming too damaged from environmental factors. Repeat this
with another six donuts and place them, again in a circle, almost on top of the first six
donuts. Continue repeating this process until there is a loop of these twine-donut
complexes as depicted in panel.

4. GENE CODING
• RNA: Ribonucleic Acid
• Before discussing the major role of DNA, it is important to discuss DNA’s first cousin, ribonucleic acid or RNA. Besides
its chemical composition, RNA has important similarities and differences with DNA. First, like DNA, RNA has four and
only four nucleotides. But unlike DNA, RNA uses the nucleotide uracil (abbreviated as U) in place 7 In addition to
physically supporting and organizing DNA, proteins in the chromosomes may also have a functional role in gene
expression. Thus, the four RNA nucleotides are adenine (A), cytosine (C), guanine (G), and uracil (U). Second, the
nucleotides in RNA also exhibit complementary base pairing. The RNA nucleotides may pair with either DNA or other
RNA molecules. When RNA pairs with DNA, G always pairs with C, 8 T in DNA always pairs with A in RNA, but A in DNA
pairs with U in RNA. When RNA pairs with RNA, then G pairs with C and A pairs with U. Third, RNA is single-stranded
while DNA is double-stranded. That is, RNA does not have the ladder-like structure of the DNA. Instead, RNA would look
like after the ladder was sawed down the middle and one half of it discarded (with, of course, the added proviso that U
would substitute for T in the remaining half). Fourth, while there is one type of DNA, there are several different types of
RNA, each of which perform different duties in the cell. Think of DNA as the monarch of the cell, giving all the orders.
Unlike human monarchs, however, king DNA is unable to leave the throne room (i.e., the cell’s nucleus) and hence, can
never execute his own orders. The different types of RNA correspond to the various types of henchmen who carry out
the King’s orders. Some occupy buildings in outlying districts (ribosomal RNA), others transport material to strategic
locations (transfer RNA), while yet others act as messengers to give instructions on what to build (messenger RNA).

4. GENE CODING

5. GENE GUN AND ALTERNATIVE TECHNIQUES
In genetic engineering, a gene gun or

biolistic particle delivery system is a device
used to deliver exogenous DNA
(transgenes), RNA, or protein to cells. By
coating particles of a heavy metal with a
gene of interest and firing these micro-
projectiles into cells using mechanical
force, an integration of desired genetic
information can be induced into cells. The
technique involved with such micro-
projectile delivery of DNA is often referred
to as biolistics.
This device is able to transform almost
any type of cell and is not limited to the
transformation of the nucleus; it can also
transform organelles, including plastids
and
2021 mitochondria Techniques in Biotechnology 33
Gene gun design
The gene gun was originally a Crosman

air pistol modified to fire dense tungsten
particles. It was invented by John C
Sanford, Ed Wolf, and Nelson Allen at
Cornell University along with Ted Klein
of DuPont between 1983 and 1986. The
original target was onions, and the
device was used to deliver particles
coated with a marker gene which would
relay a signal if proper insertion of the
DNA transcript occurred. Genetic
transformation was the demonstrated
upon observed expression of the marker
gene within onion cells.
Biolistic construct design
Biolistic transformation involves the integration of a functional fragment of DNA—known as a DNA

construct—into target cells. A gene construct is a DNA cassette containing all required regulatory
elements for proper expression within the target organism. While gene constructs may vary in their
design depending on the desired outcome of the transformation procedure, all constructs typically
contain:
Promoter:
Promoters control the location and magnitude of gene expression and function as “the steering wheel
and gas pedal” of a gene. Promoters precede the gene of interest in the DNA construct and can be
changed through laboratory design to fine-tune transgene expression. The 35S promoter from
Cauliflower mosaic virus is an example of a commonly used promoter that results in robust constitutive
gene expression within plants.

Terminator:
Terminator sequences are required for proper gene expression and are placed after the coding
region of the gene of interest within the DNA construct. A common terminator for biolistic
transformation is the NOS terminator derived from Agrobacterium tumefaciens. Due to the high
frequency of use of this terminator in genetically engineered plants, strategies have been
developed to detect its presence within the food supply to monitor for unauthorized GE crops.
Reporter gene:
A gene encoding a selectable marker is a common element within DNA constructs and is used to
select for properly transformed cells. The selectable marker chosen will depend on the species
being transformed, but it will typically be a gene granting cells a detoxification capacity for certain
herbicides or antibiotics such as kanamycin, hygromycin B, or glyphosate.

Advantages
Biolistics has proven to be a versatile method of genetic modification and it is generally preferred to
engineer transformation-resistant crops, such as cereals. Notably, Bt maize is a product of
biolistics.Plastid transformation has also seen great success with particle bombardment when
compared to other current techniques, such as Agrobacterium mediated transformation, which
have difficulty targeting the vector to and stably expressing in the chloroplast. In addition, there are
no reports of a chloroplast silencing a transgene inserted with a gene gun. Additionally, with only
one firing of a gene gun, a skilled technician can generate two transformed organisms. This
technology has even allowed for modification of specific tissues in situ, although this is likely to
damage large numbers of cells and transform only some, rather than all, cells of the tissue.

Limitations
Biolistics introduces DNA randomly into the target cells. Thus the DNA may be transformed into
whatever genomes are present in the cell, be they nuclear, mitochondrial, plasmid or any others, in
any combination, though proper construct design may mitigate this. The delivery and integration of
multiple templates of the DNA construct is a distinct possibility, resulting in potential variable
expression levels and copy numbers of the inserted gene. This is due to the ability of the constructs
to give and take genetic material from other constructs, causing some to carry no transgene and
others to carry multiple copies; the number of copies inserted depends on both how many copies of
the transgene an inserted construct has, and how many were inserted. Also, because eukaryotic
constructs rely on illegitimate recombination—a process by which the transgene is integrated into the
genome without similar genetic sequences—and not homologous recombination, they cannot be
targeted to specific locations within the genome, unless the transgene is co-delivered with genome
editing reagents.

6. BACKROSS BREEDING
In backcross method of breeding,

the hybrid and the progenies in
subsequent generations are
repeatedly backcrossed to one of
the parents. As a result, the
genotype of the backcross progeny
becomes increasingly similar to
that of the recurrent parent. The
objective of backcross method is to
improve one or two specific defects
of a high yielding variety.
6. BACKCROSS BREEDING
Pre-requisite for back cross breeding
1. A suitable recurrent parent must be available which lacks in one or
two characteristics.
2. A suitable donor parent must be available
3. The character to be transferred must have high heritability and
preferably it should be determined by one or two genes.
4. A sufficient number of back crosses should be made so that the
genotype of recurrent parent is recovered in full.

Application of back cross method
This method is commonly used to transfer disease resistance from one variety to
another. But it is also useful for transfer of other characteristics.
1. Intervarietal transfer of simply inherited characters
E.g. Disease resistance, seed coat colour
2. Intervarietal transfer of quantitative characters.
E.g. Plant height, Seed size, Seed shape.
3. Interspecific transfer of simply inherited characters
E.g. Transfer of disease resistance from related species to cultivated species.
E.g. Resistance to black arm disease in cotton from wild tetraploid species into
G.hirsutum
4. Transfer of cytoplasm
This is employed to transfer male sterility. The female parent will be having the sterile cytoplasm
and recurrent parent will be used as male parent.
E.g. Sesamum malabariucum x S.indicum
Female parent Recurrent parent.
5. Transgressive segregation
Back cross method may be modified to produce transgressive segregants. The F1 is backcrossed
to recurrent parent for 2 to 3 times for getting transgressive segregants.
6. Production of isogenic lines
7. Germplasm conversion
E.g. Production of photo insensitive line from photo Sensitive germplasm through back crossing.
This
2021
was done in the case of sorghum. Popularly known as conversion programme
Techniques in Biotechnology 42
Merits of Backcross Method
• The genotype of the new variety is nearly identical with that of the recurrent
parent, except for the genes transferred. Thus the outcome of a backcross
programme is known beforehand, and it can be reproduced any time in the future.
• It is not necessary to test the variety developed by the back cross method in
extensive yield tests because the performance of the recurrent parent is already
known. This may save upto 5 years time and a considerable expense.
• The backcross programme is not dependent upon environment, except for that
needed for the selection of the character under transfer. Therefore, off-season
nurseries and green - houses can be used to grow 2-3 generation each year. This
would drastically reduce the time required for developing the new variety.
• Much smaller population are needed in the backcross method than
in the case of pedigree method.
• Defects, such as, susceptibility to disease, of a well-adapted variety
can be removed without affecting its performance and adaptability.
Such a variety is often preferred by the farmers and the industries to an
entirely new variety because they know the recurrent variety well.
• This is the only method for interspecific gene transfers.
• It may be modified so that transgressive segregation may occur for
quantitative' characters.
Demerits of Backcross Method
1. The new variety generally cannot be superior to the recurrent parent,
except for the character that is transferred.
2. Undesirable genes closely linked with the gene being transferred may
also be transmitted to the new variety.
3. Hybridization has to be done for each backcross. This is often difficult,
time taking and costly.
4. By the time the backcross is over, the recurrent parent may have been
replaced by other varieties superior in yielding ability and other
characteristics.
7. CLONING
Cloning is the most recent evolution of
selective assisted breeding in animal
husbandry. Cloning animals is a reliable way
of reproducing superior livestock genetics
and ensuring herds are maintained at the
highest quality possible. It’s important to
remember that cloning does not manipulate
the animal’s genetic make up nor change an
animal’s DNA. It is simply another form of
assisted reproduction. Cloning allows
livestock breeders to create an exact
genetic copy of an existing animal,
essentially an identical twin. Clones are
superior breeding animals used to produce
healthier offspring.

7. CLONING

7. CLONING
• Dolly the sheep
Dolly, a Finn-Dorset ewe, was the first mammal to have been successfully
cloned from an adult somatic cell. Dolly was formed by taking a cell from the
udder of her 6-year-old biological mother. Dolly's embryo was created by
taking the cell and inserting it into a sheep ovum. It took 434 attempts before
an embryo was successful. The embryo was then placed inside a female
sheep that went through a normal pregnancy. She was cloned at the Roslin
Institute in Scotland by British scientists Sir Ian Wilmut and Keith Campbell
and lived there from her birth in 1996 until her death in 2003 when she was
six. She was born on 5 July 1996 but not announced to the world until 22
February 1997. Her stuffed remains were placed at Edinburgh's Royal
2021 Museum, part of the National Museums of Scotland.
Techniques in Biotechnology 48
7. CLONING
Cloning describes the processes used to create an exact genetic

replica of another cell, tissue or organism. The copied material, which
has the same genetic makeup as the original, is referred to as a clone.
The most famous clone was a Scottish sheep named Dolly. There are
three different types of cloning:
• Gene cloning, which creates copies of genes or segments of DNA
• Reproductive cloning, which creates copies of whole animals
• Therapeutic cloning, which creates embryonic stem cells.
Researchers hope to use these cells to grow healthy tissue to replace
injured or diseased tissues in the human body.
7. CLONING
Animal cloning offers great benefits to consumers, farmers, and endangered
species:
• Cloning allows farmers and ranchers to accelerate the reproduction of their most
productive livestock in order to better produce safe and healthy food.
• Cloning reproduces the healthiest animals, thus minimizing the use of antibiotics,
growth hormones and other chemicals.
• Consumers can benefit from cloning because meat and milk will be more
healthful, consistent, and safe. Most of the foods from cloning will be from the
offspring of clones that are not clones themselves, but sexually reproduced animals.
• Cloning can be used to protect endangered species. For example, in China,
panda cells are being kept on reserve should this species' numbers be threatened
by extinction.
7. CLONING
Ethical issues of cloning
There are a variety of ethical positions regarding the possibilities of cloning, especially human
cloning. While many of these views are religious in origin, the questions raised by cloning are faced
by secular perspectives as well. Perspectives on human cloning are theoretical, as human
therapeutic and reproductive cloning are not commercially used; animals are currently cloned in
laboratories and in livestock production.
Advocates support development of therapeutic cloning in order to generate tissues and whole
organs to treat patients who otherwise cannot obtain transplants, to avoid the need for
immunosuppressive drugs, and to stave off the effects of aging. Advocates for reproductive cloning
believe that parents who cannot otherwise procreate should have access to the technology.
Opponents of cloning have concerns that technology is not yet developed enough to be safe and
that it could be prone to abuse (leading to the generation of humans from whom organs and tissues
would be harvested), as well as concerns about how cloned individuals could integrate with families
and with society at large.
8. DNA SEQUENCING
DNA sequencing is a laboratory
technique used to determine the
exact sequence of bases (A, C,
G, and T) in a DNA molecule.
The DNA base sequence carries
the information a cell needs to
assemble protein and RNA
molecules. DNA sequence
information is important to
scientists investigating the
functions of genes.

8. DNA SEQUENCING
• What is DNA sequencing?
• Sequencing DNA means determining the order of the four chemical building blocks - called "bases" -
that make up the DNA molecule. The sequence tells scientists the kind of genetic information that is
carried in a particular DNA segment. For example, scientists can use sequence information to
determine which stretches of DNA contain genes and which stretches carry regulatory instructions,
turning genes on or off. In addition, and importantly, sequence data can highlight changes in a gene
that may cause disease.
• In the DNA double helix, the four chemical bases always bond with the same partner to form "base
pairs." Adenine (A) always pairs with thymine (T); cytosine (C) always pairs with guanine (G). This
pairing is the basis for the mechanism by which DNA molecules are copied when cells divide, and the
pairing also underlies the methods by which most DNA sequencing experiments are done. The
human genome contains about 3 billion base pairs that spell out the instructions for making and
maintaining a human being.
8. DNA SEQUENCING
• How new is DNA sequencing?
• Since the completion of the Human Genome Project, technological improvements

and automation have increased speed and lowered costs to the point where
individual genes can be sequenced routinely, and some labs can sequence well over
100,000 billion bases per year, and an entire genome can be sequenced for just a
few thousand dollars.
• Many of these new technologies were developed with support from the National
Human Genome Research Institute (NHGRI) Genome Technology Program and its
Advanced DNA Sequencing Technology awards. One of NHGRI's goals is to
promote new technologies that could eventually reduce the cost of sequencing a
human genome of even higher quality than is possible today and for less than
$1,000.
8. DNA SEQUENCING
• How new is DNA sequencing?
• Since the completion of the Human Genome Project, technological improvements

and automation have increased speed and lowered costs to the point where
individual genes can be sequenced routinely, and some labs can sequence well over
100,000 billion bases per year, and an entire genome can be sequenced for just a
few thousand dollars.
• Many of these new technologies were developed with support from the National
Human Genome Research Institute (NHGRI) Genome Technology Program and its
Advanced DNA Sequencing Technology awards. One of NHGRI's goals is to
promote new technologies that could eventually reduce the cost of sequencing a
human genome of even higher quality than is possible today and for less than
$1,000.
THANK YOU
20XX Presentation title 56

Techniques in Biotechnology: Ma. Joy O. Jocosol Eastern Samar State University

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Techniques in Biotechnology: Ma. Joy O. Jocosol Eastern Samar State University

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Chapter 2

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At the end of this topic, we are expected to:

Tissue culture, a method of

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Plant in vitro culture techniques

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Cloning is the process

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In the cloning process, the DNA is removed from cells,

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Genetic code is the term we use for the

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In genetic engineering, a gene gun or

The gene gun was originally a Crosman

Biolistic transformation involves the integration of a functional fragment of DNA—known as a DNA

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In backcross method of breeding,

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Cloning describes the processes used to create an exact genetic

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• Since the completion of the Human Genome Project, technological improvements

• Since the completion of the Human Genome Project, technological improvements

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