Pcr Mediated Gene Modification
Pcr Mediated Gene Modification - Related Videos
10:34
Identification of Sleeping Beauty Transposon Insertions in Solid Tumors using Linker-mediated PCR
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Cited by 3 •
2013
A method of identifying unknown drivers of carcinogenesis using an unbiased approach is described. The method uses the Sleeping Beauty transposon as a random mutagen directed to specific tissues. Genomic mapping of transposon insertions that drive tumor formation identifies novel oncogenes and tumor suppressor...
14:07
Genome-wide Gene Deletions in Streptococcus sanguinis by High Throughput PCR
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Cited by 9 •
2012
An efficient genome-wide single gene mutation method has been established using Streptococcus sanguinis as a model organism. This method has achieved via high throughput recombinant PCRs and transformations.
08:17
VIGS-Mediated Forward Genetics Screening for Identification of Genes Involved in Nonhost Resistance
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Cited by 22 •
2013
Virus-induced gene silencing is an useful tool for identifying genes involved in nonhost resistance of plants. We demonstrate the use of bacterial pathogens expressing GFPuv in identifying gene silenced plants susceptible to nonhost pathogens. This approach is easy, fast and facilitates large scale screening and similar protocol can be applied to studying various other plant-microbe interactions.
10:18
Agrobacterium-Mediated Virus-Induced Gene Silencing Assay In Cotton
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Cited by 159 •
2011
We present the detailed protocol for Agrobacterium-mediated virus-induced gene silencing (VIGS) assay in cotton. The tobacco rattle virus (TRV)-derived VIGS vectors were deployed to induce RNA silencing of cotton GrCLA1, Cloroplastos alterados 1 gene. The albino phenotype caused by silencing GrCLA1 was observed at the seedling stage within 2 weeks after inoculation.
02:32
Histone Modification
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2020
The histone proteins have a flexible N-terminal tail extending out from the nucleosome. These histone tails are often subjected to post-translational modifications such as acetylation, methylation, phosphorylation, and ubiquitination. Particular combinations of these modifications form “histone codes” that influence the chromatin folding and tissue-specific gene expression. Acetylation The enzyme histone acetyltransferase adds acetyl group to the histones. Another enzyme, histone deacetylase,...
FAQs
Primers act as both amplification starting points and carriers of the intended sequence change. A substitution, insertion, or deletion placed within a primer can become incorporated into the newly copied DNA strand as PCR proceeds. This design links the desired genetic alteration to the product being amplified, allowing the modified sequence to accumulate without rebuilding an entire construct.
Thermal cycling repeatedly exposes the DNA to conditions that support successive rounds of copying, while DNA polymerase extends the primer-bound strands. Each cycle creates additional templates containing the engineered sequence, so later rounds amplify molecules carrying the change. This enrichment converts an initially introduced modification into a recoverable DNA product for downstream analysis.
Compared with synthesizing an entire genetic construct, PCR-mediated gene modification focuses the change on a selected DNA molecule and uses amplification to generate the altered product. This makes the approach suitable when researchers need a defined substitution, insertion, or deletion within an existing sequence. The resulting construct can support focused studies of gene function, protein structure, or regulatory elements.
After amplification, researchers can clone the modified DNA, sequence it to examine the recovered product, and introduce it into an experimental system. These stages connect sequence engineering with biological testing: cloning provides a format for handling the altered DNA, sequencing provides information about the resulting sequence, and introduction into a system allows its effects to be examined in context.
Researchers can generate defined variants to examine how particular sequence changes affect gene function, protein structure, or regulatory elements. By comparing an altered DNA product with the relevant unmodified context in an experimental system, they can relate a designed substitution, insertion, or deletion to a biological outcome. This makes the method useful for targeted mutation analysis and functional genetics.
In genetics, the method connects a deliberately designed sequence change with analysis of its resulting DNA and biological behavior. It supports mutation analysis, recombinant DNA studies, and development of engineered genetic constructs. Because researchers can modify selected regions rather than synthesize an entire construct, the approach helps create focused variants for studying relationships between sequence, gene activity, protein properties, and regulatory control.