DNA Replication, Transcription and
Translation
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Questions
Mechanisms of DNA
replication
Transcription
Processing RNA
DNA technology. Polymerase
chain reaction. Transgenic
organisms.
Mechanisms of translation
1) Mechanisms of DNA replication
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Watson and Crick's discovery of DNA
structure in 1953 revealed a possible
mechanism for DNA replication. They
discovered that when DNA replication
occurs, it happens in a semi-
conservative fashion.
What this basically means is that each new
daugther DNA strand is made up of one
half of the original, and the other half is a
new strand.
Three proposed models of replication are conservative
replication, dispersive replication, and semiconservative
replication.
Semi-conservative model
According to the semiconservative model,
after one round of replication, every new
DNA double helix would be a hybrid that
consisted of one strand of old DNA bound to
one strand of newly synthesized DNA.
Replication fork
The point at which the two strands of
DNA are separated to allow replication
of each strand.
Let's meet the first important enzyme which make
the DNA replication happen.
DNA polymarase .There are three of these
enzymes, DNA Polymerase I,II,&III. The
one we are interested in is DNA
Polymerase III. It has the important task of
generating a new DNA strand by adding
new nucleotides.
DNA Polymerase III
Core Enzyme
DNA polymerase III is
the principal replicative
enzyme in E. coli.
DNA Polymerase III
DNA-dependent DNA polymerases are
enzymes that synthesize new DNA
strands using preexisting DNA strands
as templates
DNA Polymerase III is 15 times more
active biologically than DNA
polymerase I
DNA Polymerase III
Has two activity:
 • 5’ --> 3’ DNA polymerase activity (synthesis)
 • 3’ --> 5’ exonuclease proofreading activity
(error correction)
The 3’→5’ direction of the proof reading
exonuclease helps in the maintenance of high
replicational fidelity.
DNA polymerase III
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is able to add new nucleotides only to the
3’end (with –OH) and does not add to the 5’
end of nucleic acid.
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uses primer (small RNA) as a starting
point; the primer gives 3’ hydroxyl group
for the start of DNA-polymerase work.
DNA strand synthesis occurs only in 5’→3’
direction from the RNA primer.
Next enzymes and proteins for
synthesis
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Helicase unwinds the template strands with
breaking the hydrogen bonds
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Topoisomerase creates a small cut in the
overwounded DNA, to allow the overwounded
DNA to unwind and relax.
SSB proteins (Single Stranded Binding
proteins) is for stabilization of replication fork
Primase is for primer synthesis
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DNA-lygase
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Mechanism of synthesis
The DNA strand thus synthesized
in the 5’→3’ direction is known as
the leading strand.
Only one strand of DNA is
synthesized continuously. The
synthesis of the other strand, the
lagging strand involves a
complicated mechanism.
Mechanism of synthesis
Okazaki fragments
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The lagging strand is produced in small
fragments as Okazaki fragments. The
synthesis of the lagging strand DNA is also in
the 5’→3’ direction using a number of RNA
primers for individual stretches
RNA primers are then removed by the DNA
polymerase I having a 5’→3’ exonuclease. The
gaps are filled by this enzyme.
DNA ligase binds Okazaki fragments
2) Transcription
DNA transcription
is the synthesis of
RNA from DNA
The most important
player in prokaroytic
transcription
is RNA polymerase.
RNA polymerase
This is an enzyme whose job is to actually make
the RNA strand from the one of the DNA strands.
In the process of transcription there are
three main stages:
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Initiation: the construction of the RNA
polymerase complex on the gene's
promoter with the help of transcription
factors
Elongation: the actual transcription of
the majority of the gene into a
corresponding RNA sequence
Termination: the stop of RNA
transcription and the disassembly of the
RNA polymerase complex.
DNA template for transcription and
mechanism of process
Processing of RNA
is to generate a mRNA (for protein genes) or
a functional tRNA or rRNA from the
primary transcript.
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Types of pre-mRNA processing :
3'-Polyadenylation of mRNA: The poly-A tail
contains ~ 250 A residues in mammals, and ~ 100 in
yeasts.
5'-Capping of mRNA: the chemical structure of the
"cap" is m7G
3)Splising
DNA technology. Polymerase chain
reaction. Transgenic organisms
DNA technology has revolutionized modern
science. Recent advances in DNA technology
including cloning, PCR, recombinanat DNA
technology, DNA fingerprinting, gene therapy,
DNA microarray technology, and DNA profiling
have already begun to shape medicine, forensic
sciences, environmental sciences, and national
security.
Transgenic Organisms (genetically
modified organism)
A genetically modified organism (GMO) or genetically
engineered organism (GEO) is an organism whose
genetic material has been altered using genetic
engineering techniques.
These techniques, generally known as recombinant DNA
technology, use DNA molecules from different sources,
which are combined into one molecule to create a new set
of genes. This DNA is then transferred into an organism,
giving it modified or novel genes. Transgenic organisms,
a subset of GMOs, are organisms which have inserted
DNA that originated in a different species.
Transgenesis
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is the process of introducing an exogenous
gene – called a transgene – into a living
organism so that the organism will exhibit a
new property.
Transgenesis can be facilitated by liposomes,
plasmid vectors, viral vectors, and other.
Recombinant DNA technology
Uses of transgenic organisms:
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in toxicology: as responsive test animals (detection of
toxicants);
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in mammalian developmental genetics;
to introduce human genes into other organisms
(particularly human) for the study of disease processes;
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in molecular biology, the analysis of the regulation of gene
expression;
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in the pharmaceutical industry,
in biotechnology: as producers of specific proteins;
genetically engineered hormones to increase milk yield,
meat production; agriculture (food production)
POLYMERASE CHAIN REACTION
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The polymerase chain reaction
(PCR) was discovered by Kary
Mullis.
He won a Nobel Prize in 1993 and
became very "FAMOUS" because of
this discovery.
PCR is the method of DNA
ampliphication (or increasing of
quantity of DNA molecules).
This figure represents the PCR
reaction.
DNA profiling (DNA testing)
is used in establishing proof of paternity
or identifying siblings. While DNA
contains material common to all humans,
some portions are unique to each
individual. DNA testing can help solve
crimes by comparing the DNA profiles of
suspects to offender samples.
DNA Paternity Test
One of the most popular and readily available of
these tests is the DNA paternity test. During this
test the genetic profile of the child is compared to
the genetic profile of the alleged father, and if a
large enough number of similarities are found,
then it can be concluded that the alleged father is
indeed the biological father of the child.
The accuracy of determining is 99.99%
Translation
is the synthetic of proteins
molecules on RNA template
Stages:
- Initiation
- Elongation
- Termination