Gene-targeted Mutagenesis
Gene-targeted Mutagenesis - Related Videos
08:48
Targeted in Situ Mutagenesis of Histone Genes in Budding Yeast
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Cited by 5 •
2017
A strategy for generating mutations in histone genes at their endogenous location in Saccharomyces cerevisiae is presented.
07:25
Transfection and Mutagenesis of Target Genes in Mosquito Cells by Locked Nucleic Acid-modified Oligonucleotides
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Cited by 3 •
2010
Oligonucleotides can be used to site specifically substitute a single nucleotide of transfected target genes in both Anopheles gambiae and Anopheles stephensi cells.
08:56
Recombineering and Gene Targeting
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2023
One of the most widely used tools in modern biology is molecular cloning with restriction enzymes, which create compatible ends between DNA fragments that allow them to be joined together. However, this technique has certain restrictions that limit its applicability for large or complex DNA construct generation. A newer technique that addresses some of these shortcomings is recombineering, which modifies DNA using homologous recombination (HR), the exchange between different DNA molecules based...
02:29
In-vitro Mutagenesis (Gene Knockouts)
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2026
To learn more about the function of a gene, researchers can observe what happens when the gene is inactivated or “knocked out,” by creating genetically engineered knockout animals. Knockout mice have been particularly useful as models for human diseases such as cancer, Parkinson’s disease, and diabetes. The Process Genes can be randomly knocked out, or specific genes can be targeted. To knock out a particular gene, an engineered piece of DNA called a targeting vector is used to replace the...
07:18
Gene-targeted Random Mutagenesis to Select Heterochromatin-destabilizing Proteasome Mutants in Fission Yeast
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Cited by 1 •
2018
This article describes a detailed methodology for random mutagenesis of a target gene in fission yeast. As an example, we target rpt4+, which encodes a subunit of the 19S proteasome, and screen for mutations that destabilize heterochromatin.
FAQs
After a chosen locus is modified, the cell’s DNA-repair response determines the resulting allele. Repair may create a small insertion or deletion, which can disrupt gene activity, or use a donor template to replace a sequence. Thus, the same targeting strategy can produce either a gene-disrupting change or a more defined sequence replacement, depending on the repair route engaged.
Gene-targeted mutagenesis relies on two complementary targeting approaches: a nuclease can act at the selected locus, whereas a recombination system can direct genetic exchange at that site. Their shared purpose is positional precision, but the resulting change depends on whether the experiment is designed to disrupt an existing sequence or introduce a replacement sequence.
Choosing a defined locus is essential because interpretation depends on connecting one intentional DNA change with one observed trait. When the altered sequence is linked to a change in an organism, cell, or biological trait, researchers can use that relationship to clarify gene function. This locus-to-phenotype connection is central to functional genetic analysis.
A donor template supports sequence replacement rather than relying only on a small insertion or deletion. In a targeting experiment, researchers direct the editing or recombination system to the chosen locus and use the intended sequence as the replacement source. The resulting change can help examine a defined genetic alteration, including a disease-associated variant.
The main experimental outcome is a relationship between a specific engineered mutation and an observable phenotype. If altering the locus changes a trait, the result can support a role for that gene in the trait or process under study. Such evidence is used for functional analysis and for validating whether a disease-associated variant produces a biologically relevant effect.
Beyond basic gene-function studies, the approach supports several genetics applications. Researchers can create experimental models carrying specific genetic traits, investigate mechanisms associated with disease, and study developmental effects. The same strategy can also contribute to evaluating potential therapeutic strategies, because defined sequence changes provide a way to examine how altering a gene influences a biological outcome.