Cyagen’s innovative TurboKnockout® technology is an advanced version of ES cell-mediated gene targeting, which has shortened the construction turnaround time from one year to as few as six months – on par with CRISPR. Compared to CRISPR/Cas9, TurboKnockout® provides accurate genetic changes without off-target effects and can perform complex genetic modifications – including large regions up to 300 kb.
The TurboKnockout® gene targeting service can be used for complex gene modeling projects to provide C57BL/6 or BALB/c mouse models with accurate genetic modification of large genomic regions. This proprietary gene targeting method eliminates at least two generations of breeding, shortening production time by 4-6 months as compared to the industry standard.
What is Recombinase-Mediated Cassette Exchange (RMCE) and BAC?
Recombinase-mediated cassette exchange (RMCE) implements site-specific recombination to achieve systematic, repeated modification of higher eukaryotic genomes. The recombinase-mediated cassette exchange (RMCE) approach remains a powerful tool for the insertion of large gene fragments, and may be used to efficiently generate conditional, reporter, and transgenic mouse models. RMCE provides highly efficient transfer of genes of interest (GOIs) and is readily adapted to generate humanized mouse models – reproducing the human expression patterns in mice.
Bacterial artificial chromosome (BAC) is a low-copy vector that can hold more than 300 kb of foreign DNA. Through BAC recombination, larger genes and regulatory sequences can be introduced - which is closer to the expression pattern of endogenous genes.
TurboKnockout®, RMCE, and BAC: Accelerating Antibody Discovery Research
Based on TurboKnockout® (ES) cell targeting techniques, we have used RMCE-based techniques (with BAC fusion and 3 targeting rounds) to achieve large-fragment knockin (LFKI) humanization and genetic modifications for regions up to 300 kb. Additionally, multi-step BAC reorganization at the ES cell level can be used to shorten the production cycle.
CRISPR/Cas9 Versus TurboKnockout®
TurboKnockout Advantages
1. Freedom to Operate: No Risk of Patent Infringement
Compared with CRISPR/Cas9 techniques, TurboKnockout® is free of patent disputes and is the technique of choice for new drug development projects.
2. Success Rate: 100% guaranteed germline transmission (GLT)
TurboKnockout® is based on traditional ES cell targeting techniques; it can be used for complex gene knockouts for mice, providing models with accurate genetic modification, 100% germline transmission, and no off-target effects.
3. Shorter Turnaround: Founders as fast as 6 months
Innovative techniques eliminate two generations of breeding & shorten production time by 4-6 months:
a) Super competent ES cell line generates 100% ESC-derived founders, avoiding the chimera phase;
b) Self-removing Neo selection cassette that circumvents the need to breed to Flp deleter mice.
For conditional knockouts (cKO, floxed), it may be possible to breed founders directly to your tissue-specific Cre mice.
4. Large Fragment Knockin: Insert fragments up to 300 kb
Limited by the capacity of DNA cloning vectors, foreign gene fragments integrated into genetically modified animals are usually less than 30kb. At this size limitation, key elements in regulating gene activity are often lost. BAC can accommodate foreign DNA of more than 300 kb. By introducing larger genes and regulatory sequences through BAC modification, this allows greater control over the expression pattern of genes.
5. Flexible Floxing: No size limitations for your floxed region
With the advantages of simple operation and high recombination rate, the Cre-Lox system has become a powerful tool for genetic manipulation across in vivo and in vitro models. The Cre-Lox system enables gene expression with exceptional spatial (gene expression in specific cells, tissues, or entire organisms) and temporal (at specific time points through the application of the Cre-Lox system) control.
6. 100% Money-Back Guarantee
To learn more about TurboKnockout® Technology, please visit our page.
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