Polymerase Chain Reaction PPT

7/28/2019 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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Polymerase Chain Reaction PPT