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Polymerase chain reaction

life science Maturity 9-11 Next article →

The polymerase chain reaction, or PCR, is a vital laboratory method used to amplify specific DNA sequences. This process allows scientists to take a very tiny amount of genetic material and make millions of copies of it.

Polymerase chain reaction-en
Polymerase chain reaction-en

By creating these large quantities, researchers can perform detailed studies that would otherwise be impossible. PCR is now a fundamental tool in many different scientific fields. It is used for medical research, forensic science, and the study of ancient DNA.

US Army CID agents at crime scene
US Army CID agents at crime scene

Without this technology, many biological samples would remain too small to analyze effectively.

The mechanism of PCR relies on a process called thermal cycling. This involves exposing a reaction mixture to repeated cycles of heating and cooling.

PCR masina kasutamine
PCR masina kasutamine

Each cycle consists of three main temperature-dependent steps: denaturation, annealing, and extension. During denaturation, the reaction is heated to a high temperature, often around 94–98°C. This high heat breaks the hydrogen bonds between the two strands of the DNA double helix. As a result, the two strands separate into single molecules. This provides the necessary templates for the next steps of the reaction.

Once the strands are separated, the temperature is lowered during the annealing step. This cooling allows primers to bind to the single-stranded DNA. Primers are short, single-stranded DNA fragments known as oligonucleotides. They are designed to be complementary to the specific target region of the DNA.

Primers RevComp
Primers RevComp

It is critical to choose the correct temperature for this step. If the temperature is too low, the primers might bind to the wrong places. If the temperature is too high, the primers will not bind at all. Successful annealing creates a double-stranded region where the enzyme can begin its work.

The third step is the extension or elongation phase. During this stage, the temperature is raised to an optimal level for the DNA polymerase enzyme. This enzyme assembles a new DNA strand by adding free nucleotides, called dNTPs, to the primer. These dNTPs are the essential building blocks of DNA.

Polymerase chain reaction-en
Polymerase chain reaction-en

The polymerase moves along the template strand, creating a complementary sequence. As the reaction progresses, the newly generated DNA strands themselves become templates for the next cycle. This creates a chain reaction that results in exponential amplification.

Most modern PCR methods utilize a specific type of enzyme called Taq polymerase. This is a thermostable DNA polymerase, meaning it can withstand high temperatures without breaking down. It was originally isolated from a thermophilic bacterium named Thermus aquaticus.

Primitive PCR machine for scrap
Primitive PCR machine for scrap

Before Taq polymerase was used, scientists had to manually add new enzymes during every single cycle. This was a very tedious and expensive process. Using a heat-stable enzyme makes the process much faster and more efficient. Some scientists also use Pfu polymerase, which is slower but has higher accuracy because it can proofread its work.

PCR was invented in 1983 by the American biochemist Kary Mullis. He developed this method while working at the Cetus Corporation.

His work, along with the DNA manipulation techniques developed by Michael Smith, changed biology forever. In 1993, Mullis and Smith were jointly awarded the Nobel Prize in Chemistry. This recognition highlighted the massive impact of their discovery on the scientific community.

The power of PCR lies in its ability to produce massive amounts of DNA through geometric growth. In each cycle, the number of DNA target sequences is doubled.

Exponential Amplification
Exponential Amplification

If a reaction is highly efficient, a single copy of DNA can increase to one billion copies after just 30 cycles. This rapid increase is calculated using the formula 2^n, where n is the number of cycles. This level of amplification is essential for practical applications. It allows for the detection of pathogens in infectious diseases and the analysis of genetic fingerprints for parentage testing.

Pcr fingerprint
Pcr fingerprint

It also enables the construction of DNA-based phylogenies to study the relationships between different living things.

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🖼️ Images & Media (11)
File:PCR tubes.png
PCR tubes.png
File:PCR masina kasutamine.jpg
PCR masina kasutamine.jpg
File:Primitive PCR machine for scrap.JPG
Primitive PCR machine for scrap.JPG
File:Polymerase chain reaction-en.svg
Polymerase chain reaction-en.svg
File:Roland Gel.JPG
Roland Gel.JPG
File:Tucker PCR.png
Tucker PCR.png
File:Exponential Amplification.svg
Exponential Amplification.svg
File:Pcr fingerprint.png
Pcr fingerprint.png
File:US Army CID agents at crime scene.jpg
US Army CID agents at crime scene.jpg
File:Primers RevComp.svg
Primers RevComp.svg
File:Baby Blue - a prototype polymerase chain reaction (PCR), c 1986. (9663810586).jpg
Baby Blue - a prototype polymerase chain...
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