polymerase chain reaction ppt
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Polymerase Chain
Reaction - PCR
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The polymerase chain reaction
(PCR) is a technique to amplify apiece of DNA very rapidly outside
of a cell.
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Some applications of PCR.
Forensic medicine.
Preimplantation Genetic Diagnosis
(PGD).Archeology.
Paternity testing.
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The polymerase chain reaction
(PCR).
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A cycle of PCR consists of three steps.
DNA denaturation at 95 degrees C.
Primer annealing at 50-60 degrees C.
DNA polymerization by a thermostableDNA polymerase at 72 degrees C.
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Starting with a single molecule of
DNA, 25 rounds or cycles of PCRwill produce about 10 million
identical DNA molecules!!
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Forensic uses of PCR
PCR can be used to amplify DNA from a
small amount of cells (about 1000 cells).
The amplified DNA from cells can beused in DNA fingerprinting analysis to
determine who was at the crime scene.
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DNA fingerprinting using PCR in forensic investigations.
DNA is isolated from blood at a crime
scene and amplified by PCR.
The amplified DNA is digested withrestriction enzymes and resolved on an
agarose gel.
Southern blot analysis is performed to
give a DNA fingerprint.
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Restriction fragment analysis by
Southern blotting.
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DNA fingerprints from a murder case.
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Individuals have unique DNAfingerprints because of restriction
length polymorphisms (RFLPs).
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How reliable is DNA fingerprinting?
DNA regions chosen are ones known to be
highly variable from one person to another.
In most forensic cases, the probability of two
people having identical DNA fingerprints is
between one chance in 100,000 and one in a
billion.
The exact number depends on the number of
probes used to different regions of humanchromosomal DNA.
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Many argue that DNA evidence is
more reliable than eyewitnesses
in placing a suspect at the scene
of a crime.
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Satellite DNA can be used as markers for DNA
fingerprinting.
Satellite DNA consists of tandemly repeated
base sequences within the human genome.
The most useful satellite DNA for forensic
purposes are microsatellites having repeating
units of only a few base pairs, and the number
of repeats are highly variable from one person
to another.
Microsatellite DNA is also called a simpletandem repeats (STRs).
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An example of simple tandem
repeat (STR) alleles.
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STRs in DNA fingerprinting.
The greater the number of STRsanalyzed in a DNA sample, the morelikely the DNA fingerprint is unique to anindividual.
PCR is used to selectively amplifyparticular STRs before electrophoresis.
PCR is especially valuable when DNA isin poor condition or available in minutequantities.
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PCR use in Pre-implantation Genetic
Diagnosis (PGD).
PGD is a way to determine if human embryosfrom in vitro fertilization have genetic defects(for example, cystic fibrosis).
A cell is removed from an eight cell embryo andthe DNA is analyzed by PCR for geneticdefects.
Only healthy embryos are implanted into amothers uterus.
Should this technology be used for things likegender selection?
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