Related Experiment Video
Updated: Jan 25, 2026
07:06
Principles of Site-Specific Recombinase SSR Technology
Published on: May 29, 2008
21.5K
High-resolution specificity profiling and off-target prediction for site-specific DNA recombinases
Jeffrey L Bessen1,2,3, Lena K Afeyan1,2,3, Vlado Dančík4
1Merkin Institute of Transformative Technologies in Healthcare, Broad Institute of Harvard and MIT, Cambridge, MA, 02142, USA.
Nature Communications
|April 28, 2019
Summary
Rec-seq is a new method that reveals DNA specificities of site-specific recombinases (SSRs). This tool accurately identifies potential off-target sites for genome editing applications.
Area of Science:
- Molecular Biology
- Genetics
- Biotechnology
Background:
- Site-specific recombinases (SSRs) are valuable genome editing tools.
- Altering SSR DNA specificity is challenging, limiting their application.
- Understanding SSR specificity is crucial for precise genome engineering.
Purpose of the Study:
- To develop a method for comprehensive and unbiased characterization of SSR DNA specificities.
- To identify DNA specificity determinants and potential off-target substrates of SSRs.
- To enable the rational design and improvement of SSRs for genome editing.
Main Methods:
- Rec-seq: a novel method for high-resolution DNA specificity profiling of SSRs.
- Application of Rec-seq to natural and evolved SSRs, including Cre recombinase variants.
- In silico prediction and experimental validation of off-target recombination events.
Main Results:
- Rec-seq successfully identified DNA specificity determinants for various SSRs.
- Uncharacterized SSR-DNA interactions were revealed, some not predicted by structural data.
- Rec-seq predicted and validated off-target recombination by Tre and Brec1 in human cells.
Conclusions:
- Rec-seq provides high-resolution, single-nucleotide characterization of SSR DNA specificity.
- The method facilitates rapid assessment and informs the development of novel SSRs.
- Rec-seq is a powerful tool for understanding and engineering SSRs for precise genome editing.
Related Concept Videos
01:13Sensitivity, Specificity, and Predicted Value
1.2K
In healthcare diagnostics, laboratory tests play a crucial role in identifying and diagnosing a wide range of medical conditions. However, interpreting test results is not always straightforward. An abnormal test result does not always confirm the presence of a disease, just as a normal result does not guarantee its absence. To assess the reliability of these diagnostic tools, healthcare practitioners rely on two key statistical indicators: sensitivity and specificity.
Sensitivity is the...
Sensitivity is the...
1.2K
02:53Conservative Site-specific Recombination and Phase Variation
6.7K
Because the DNA segments are cut and reorganized in a direction-specific manner, site-specific recombination has emerged as an efficient genetic engineering technique. Flippase and Cyclization recombinases or Flp and Cre, respectively, are two members of the tyrosine recombinase family derived from bacteriophages, that are used to mediate site-specific DNA insertions, deletions, and targeted expression of proteins in mammalian cell lines.
The recognition sites for Cre recombinase called LoxP...
The recognition sites for Cre recombinase called LoxP...
6.7K
01:16Specific Heat
67.3K
The specific heat capacity of a substance refers to the energy required to increase the temperature of one gram of that substance by one degree Celcius. Specific heat capacity is often represented in calories (cal), grams (g), and degrees Celsius (oC), but can also be expressed in joules (J), kilograms (kg), and Kelvin (K), among other units.
For example, increasing the temperature of one gram of water by 1°C requires one calorie of heat energy and can be written as 1 cal/g-°C, or...
For example, increasing the temperature of one gram of water by 1°C requires one calorie of heat energy and can be written as 1 cal/g-°C, or...
67.3K
01:58Cell Specific Gene Expression
16.3K
Multicellular organisms contain a variety of structurally and functionally distinct cell types, but the DNA in all the cells originated from the same parent cells. The differences in the cells can be attributed to the differential gene expression. Liver cells, whose functions include detoxification of blood, production of bile to metabolize fats, and synthesis of proteins essential for metabolism, must express a specific set of genes to perform their functions. Gene expression also varies with...
16.3K
01:58Cell Specific Gene Expression
5.4K
5.4K
01:27Density, Specific Weight, Specific Gravity and Compressibility of Fluid
1.6K
Density, specific weight, specific gravity, and compressibility are fundamental properties of fluids. Density is the mass per unit volume, characterizing the mass of a fluid system. It influences buoyancy, pressure, flow dynamics, viscosity, thermal conductivity, and sound propagation. For instance, in pipeline design, accurate density measurements ensure that the pipeline can handle the fluid's mass.
Specific weight represents the weight per unit volume and is calculated by multiplying...
Specific weight represents the weight per unit volume and is calculated by multiplying...
1.6K