When do PCR Tests Work Best?

 

Scientists and researchers rely on PCR (polymerase chain reaction) tests to detect various infectious diseases and genetic changes. By conducting these tests, scientists can find the DNA or RNA of a pathogen. A PCR test is a preferred option when the root cause of the disease is a virus or bacteria. However, there are certain circumstances when PCR tests work best. It’s the reason why they are conducted in a PCR workstation.

Purpose of PCR tests

The main objectives of conducting PCR tests are listed below:

1. Determine infectious diseases
2. Detect a genetic change that can do harm
3. Identify cancer cells that may be overlooked in other cells
4. Gene mapping

Types of PCR tests

A PCR test can be of various types as listed below:

1. Real-time PCR: Here, a fluorescent reporter is used to detect the DNA amplification in real-time.
2. Multiplex PCR: A multiplex PCR test is implemented for the amplification of several targets.
3. Nested PCR: A nested PCR curbs the non-specific binding of products and enhances sensitivity and specificity.
4. Quantitative PCR: A quantitative PCR test is grounded on the DNA amplification linearity. This PCR test is used to identify and quantify a known sequence in the sample.
5. Arbitrary Primed PCR: This DNA fingerprinting technique is embedded in PCR. It implements primers with the DNA sequence chosen arbitrarily.

How can you set up a PCR test?

Setting up a PCR test is never easy as scientists need to get hold of the following elements:

1. The DNA template that has to be copied
2. DNA nucleotide bases as they are the building blocks of DNA. Researchers and molecular biologists require these bases to develop the new strand of DNA.
3. Primers (short stretches of DNA) that start the PCR reaction
4. Taq polymerase enzyme to build the new DNA bases
5. Make sure the conditions are perfect for the reaction by buffering.

Researchers get all the necessary arrangements in a PCR workstation. For this reason, a PCR test is always carried out at this workspace.

Steps involved in PCR tests

Given below is the list of steps involved in a PCR test:

1. DNA isolation: Being a researcher, no matter whether you are isolating bacterial plasmids or genomic DNA, you can’t bypass this crucial first step. During this phase, certain nucleic acid purification products are picked to give maximum yield, integrity, and purity from any type of sample.
2. Primer design: You have to choose high-quality primers with good designs. The length of the primers will be between 18-30 nucleotides. And yes, always remember to keep the melting temperatures between 65⁰C and 75⁰
3. Enzyme selection: Now, your task is to select the enzyme. In 99.9% of cases, DNA polymerase is used since this compound plays a great hand in synthesizing new strands of DNA.
4. Thermal cycling: A PCR comprises heating and cooling cycles for the purpose of DNA amplification. Hence, choosing the right thermal cycler will determine the success or failure of a PCR reaction.
5. PCR plastics: You need to use the apt PCR plastics for your experiment. If you choose the right plastic, it will enhance the authenticity of your PCR outcomes. Besides this aspect, the perfect fit and uniform heat transfer during the thermal cycling process are critical to productive PCR.
6. Amplicon analysis: Molecular biologists implement the nucleic acid electrophoresis techniques to detect, differentiate, quantify, and purify amplicons. However, they have to select the right instruments for this technique to give accurate results.

Wrapping up

Conducting a PCR test is an accurate method for detecting multiple infectious diseases. With this test, researchers and pathologists can make diagnoses before symptoms occur. More often, forensic researchers also rely on the results of a PCR test to identify a criminal, as this test helps in evaluating the gene expression.

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