Amplicon Pcr
Amplicon Pcr - Related Videos
03:41
Agarose Gel Electrophoresis of DNA Amplicons Post PCR: A Method to Analyze Products of Multiplex PCR and Evaluate PCR Reaction Success
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2025
In this video, we demonstrate the separation of bacterial PCR-amplified DNA using agarose gel electrophoresis. Agarose gel functions as a molecular sieve, enabling the negatively-charged DNA to migrate based upon their size under an applied electric field.
08:57
Amplicon Sequencing using the Long-Read Sequencing Technologies
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2025
This protocol was optimized for targeted deep sequencing of 18 drug-resistance regions in Mycobacterium tuberculosis using a long-read sequencing platform, followed by analysis with a tuberculosis-specific bioinformatics pipeline designed for long-read data.
10:24
Next-generation Sequencing of 16S Ribosomal RNA Gene Amplicons
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Cited by 119 •
2014
Characterizing microbial community has been a longstanding goal in environmental microbiology. Next-generation sequencing methods now allow for the characterization of microbial communities at an unprecedented depth with minimal cost and labor. We detail here our approach to sequence bacterial 16S ribosomal RNA genes using a benchtop sequencer.
13:27
PCR: The Polymerase Chain Reaction
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2023
The polymerase chain reaction, or PCR, is a technique used to amplify DNA through thermocycling – cyles of temperature changes at fixed time intervals. Using a thermostable DNA polymerase, PCR can create numerous copies of DNA from DNA building blocks called dNTPs. There are three steps in PCR: denaturation, annealing, and elongation. Denaturation is the first step in the cycle and causes the DNA to melt by disrupting hydrogen bonds between the bases resulting in single-stranded DNA. Annealing...
03:29
Capillary Electrophoresis: A Technique to Analyze Fluorescently Labeled Polymerase Chain Reaction Amplicons
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2025
This video describes the technique of capillary electrophoresis to analyze fluorescently labeled PCR amplicons obtained from various samples based on their size. It generates a migratory pattern that determines the fragments’ base-pair lengths with the help of the labeled fluorescent dye in each fragment.
FAQs
Sequence-specific primers guide amplification toward a defined DNA locus rather than the entire sample. Their selection establishes the boundaries of the target region that becomes the amplicon. During repeated thermal cycles, primer annealing positions the primers for DNA polymerase-mediated extension, giving the reaction its targeted character and producing material suitable for later analysis.
Repeated cycles increase the amount of the selected target sequence because each cycle approximately doubles the amplicon. This cumulative amplification converts a small starting quantity into many copies that can be examined more readily. The resulting sensitivity supports applications in which the original sample contains limited DNA but a defined region must still be detected or analyzed.
The three recurring stages provide the sequence of events needed to copy the target region. Denaturation, primer annealing, and DNA polymerase-mediated extension occur in repeated thermal cycles, with each stage contributing to formation of additional amplicons. Together, they enable selective, progressive accumulation of the defined DNA sequence rather than an undirected increase in all sample DNA.
Targeting a defined locus concentrates the experiment on a selected genetic region, which makes the resulting material suited to focused analysis. This approach can reveal genetic variants or support identification of pathogen-associated target sequences, while also providing a defined region for sequencing. The method therefore emphasizes depth at a chosen locus rather than broad analysis across unspecified DNA.
A basic workflow selects the DNA region of interest, uses sequence-specific primers, and subjects the reaction to repeated thermal cycles. Those cycles alternate denaturation and primer annealing with DNA polymerase-mediated extension. As amplification proceeds, the defined target accumulates as an amplicon, which can then be analyzed for variants, pathogen detection, construct verification, or sequencing preparation.
Researchers use the technique when they need targeted information from a particular DNA region. In genetics, it helps identify variants; in microbiology and diagnostics, it supports pathogen detection. It can verify engineered constructs and prepare selected regions for sequencing. These uses also extend to evolutionary studies, where focused sequence analysis can contribute to comparisons among biological samples.