Whole Genome Amplification
Whole Genome Amplification - Related Videos
09:31
Amplification, Next-generation Sequencing, and Genomic DNA Mapping of Retroviral Integration Sites
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Cited by 56 •
2016
We describe a protocol for amplifying retroviral integration sites from the genomic DNA of infected cells, sequencing the amplified virus-host junctions, and then mapping these sequences to a reference genome. We also describe techniques to quantify the distribution of integration sites relative to various genomic annotations using BEDTools.
08:31
Isolation and Genome Analysis of Single Virions using 'Single Virus Genomics'
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Cited by 2 •
2013
Single Virus Genomics (SVG) is a method to isolate and amplify the genomes of single virons. Viral suspensions of a mixed assemblage are sorted using flow cytometry onto a microscope slide with discrete wells containing agarose, thereby capturing the virion and reducing genome shearing during downstream processing. Whole genome amplification is achieved using multiple displacement amplification (MDA) resulting in genomic material that is suitable for sequencing.
09:44
Rapid Amplification of cDNA Ends
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2023
Source: Pablo Sanchez Bosch2, Sean Corcoran2 and Katja Brückner1,2,3 1Eli and Edythe Broad Center of Regeneration Medicine and Stem Cell Research 2Department of Cell and Tissue Biology, 3Cardiovascular Research Institute, University of California San Francisco, San Francisco, CA, USA Rapid Amplification of cDNA Ends (RACE) is a technique that allows amplification of full-length cDNA from mRNA by extending to the 3’ or 5’ end, even without prior knowledge of the sequence (Frohman et al.,...
08:31
Genome Editing
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2023
A well-established technique for modifying specific sequences in the genome is gene targeting by homologous recombination, but this method can be laborious and only works in certain organisms. Recent advances have led to the development of “genome editing”, which works by inducing double-strand breaks in DNA using engineered nuclease enzymes guided to target genomic sites by either proteins or RNAs that recognize specific sequences. When a cell attempts to repair this damage, mutations can be...
14:26
Genome-wide Purification of Extrachromosomal Circular DNA from Eukaryotic Cells
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Cited by 59 •
2016
This paper presents a sensitive method called Circle-Seq for purifying extrachromosomal circular DNA (eccDNA). The method encompasses column purification, removal of remaining linear chromosomal DNA, rolling-circle amplification and high-throughput sequencing. Circle-Seq is applicable to genome-scale screening of eukaryotic eccDNA and studying genome instability and copy-number variation.
FAQs
Multiple displacement amplification begins when random primers bind at many positions across the available template. Phi29 DNA polymerase then extends those primers and displaces previously synthesized strands, producing long DNA products while the reaction remains isothermal, meaning it does not rely on repeated temperature cycling. This combination enables broad genome copying from limited starting material for downstream analysis.
Whole Genome Amplification can be performed through more than one amplification strategy. Multiple displacement amplification uses phi29 polymerase under isothermal conditions, whereas other approaches use PCR cycling. That distinction matters when interpreting the resulting material because the selected method determines how DNA synthesis is initiated and carried out, while both approaches serve the broader goal of enabling genomic analysis from scarce samples.
Amplification bias can make some genomic regions more strongly represented than others, so the amplified material may not reflect the starting genome uniformly. Contamination is another concern when very little original DNA is present, because unwanted DNA can influence downstream sequencing or genetic analysis. Careful controls are therefore essential for distinguishing sample-derived findings from technical artifacts.
An analysis workflow starts with a limited clinical, environmental, cellular, or microbial specimen, followed by selection of a WGA strategy and amplification of its available DNA. The resulting material can then be directed to sequencing or other genetic analysis. Because the input is scarce, interpreting results requires attention to the method’s potential bias and to contamination controls.
When a pathogen cannot readily be characterized from abundant material, Whole Genome Amplification can provide DNA for genomic investigation from an uncultured microbe or a limited clinical specimen. The amplified product may support pathogen genome characterization and sequencing, while environmental samples extend the approach beyond clinical settings. Its value is greatest when the original biological material is insufficient for direct analysis.
In immunology and infection research, the method can connect scarce biological material with questions about both the infectious agent and the host. Researchers can use it to examine pathogen genomes, investigate host genetic variation, or study rare infected cells. These applications expand analysis to samples that might otherwise provide too little DNA, but findings still require careful interpretation because amplification can introduce bias.