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6 rDNA Technology

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34 views58 pages

6 rDNA Technology

Uploaded by

Nikhil Kumar
Copyright
© © All Rights Reserved
We take content rights seriously. If you suspect this is your content, claim it here.
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Download as PPTX, PDF, TXT or read online on Scribd
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Recombinant DNA technology

• Recombinant DNA technology comprises altering


genetic material outside an organism to obtain
enhanced and desired characteristics in living
organisms or as their products.

• Recombinant DNA technology was invented largely through the work of


American biochemists Stanley N. Cohen, Herbert W. Boyer, and Paul Berg.

• The recombinant DNA technology emerged with the discovery


of restriction enzymes in the year 1968 by Swiss
microbiologist Werner Arber.
• Process of Recombinant DNA Technology
• The complete process of recombinant DNA technology includes multiple steps,
maintained in a specific sequence to generate the desired product.
• Step-1. Isolation of Genetic Material.
• The first and the initial step in Recombinant DNA technology is to isolate the
desired DNA in its pure form i.e. free from other macromolecules.
• Step-2.Cutting the gene at the recognition sites.
• The restriction enzymes play a major role in determining the location at which
the desired gene is inserted into the vector genome. These reactions are called
‘restriction enzyme digestions’.
• Step-3. Amplifying the gene copies through Polymerase chain reaction (PCR).
• It is a process to amplify a single copy of DNA into thousands to millions of
copies once the proper gene of interest has been cut using restriction enzymes.
• Step-4. Ligation of DNA Molecules.
• In this step of Ligation, the joining of the two pieces – a cut fragment of DNA and
the vector together with the help of the enzyme DNA ligase.
• Step-5. Insertion of Recombinant DNA Into Host.
Common sources for DNA isolation

Whole blood ,
Hair,
Bones,
Nails,
Tissues,
Blood stains,
Saliva ,
Buccal (cheek) swabs,
Epithelial cells,
Bacteria,
Animal tissues, or
Plants
In a commercial kit
the common lysis
solutions contain:
sodium chloride;
tromethamine (also
known as Tris), which
is a buffer to retain
constant pH;
ethylenediaminetetraa
cetic acid (EDTA),
which binds metal
ions; and sodium
dodecyl sulfate (SDS),
which is a detergent.
A common enzyme
used in DNA
extraction is
Proteinase K
• DNA LIGATION
• DNA Amplification/DNA Cloning
• GEL ELECTROPHORESIS
•
• Cloning Libraries
• It is a collection of cloned DNA fragments that includes the
gene of interest.
• A DNA library generally is stored in a population of
bacterial cells.
• There are two types of DNA libraries:
• genomic and cDNA.
• A genomic library is a collection of DNA fragments contained
within self-replicating vectors that represent the entire
genome of the individual from which the DNA was made.
• The cDNA library represents a collection of only those DNA
fragments that were transcribed into mRNA in the cell from
which the mRNA was isolated.
• PCR/ Polymerase Chain reaction
• Nucleic Acid Hybridization
• Nucleic Acid Hybridization Base pairing permits the detection of a sequence that is
complementary to the probe.
• Microarray- Detection Method of Genes
• Genetically Modified Organism (Transgenic
Organism)
• In GMO, genetic material (DNA) is altered or artificially
introduced using genetic engineering techniques.
• Genetic modification involves the mutation, insertion, or
deletion of genes.
• Inserted genes usually come from a different organism (e.g.
In Bt cotton, Bt genes from bacterium Bacillus
thuringiensis are induced).
• Genetic modification is done to induce a desirable new
trait which does not occur naturally in the species.
• Major GM Crops
• Bt Cotton
• Bt cotton is insect-resistant cotton variety.
• Strains of the bacterium Bacillus
thuringiensis produce different Bt toxins.
• Bt toxins are insecticidal to the larvae of moths,
bollworms, etc. but are harmless to other forms of life.
• In 2002, a joint venture between Monsanto and
Mahyco introduced Bt cotton to India.
• Major GM Crops
• Golden rice
• Golden rice is a variety of rice (Oryza
sativa) produced to biosynthesize beta-
carotene, a precursor of Vitamin A, in the
edible parts of rice.
• It is mostly consumed in areas with a shortage
of dietary vitamin A.
• Major GM Crops
• Bt Brinjal
• Brinjal is India’s second most consumed
vegetable after potatoes.
• Bt brinjal is created by inserting a crystal
protein gene from the soil bacterium Bacillus
thuringiensis.
• The Bt brinjal has been developed to give
resistance to the Brinjal Fruit and Shoot
Borer (FSB).
• Transgenesis is the process of introduction of extra
(foreign/exogenous) DNA into the genome of the animals to
create and maintain stable heritable characters.
•
• The foreign DNA that is introduced is called the transgene and
the animals that are produced by DNA manipulations are
called transgenic animals or the genetically engineered or
genetically modified organisms.
• GENE THERAPY

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