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By combining multiplex targeting of yellow with the introduction of an ssODN donor template containing exogenous sequence, we successfully replaced yellow with an attP docking site.
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Furthermore, multiplex targeting enables deletion of 133 Kb, accounting for approximately 3% of the entire E. coli genome.
The easily implementable and modular nature of our of CRISPR plasmid system enables multiplex targeting.
Multiplex targeting has always been considered to be an advantage of the CRISPR/Cas9 systems17,18; however, it should be noted that multiplex targeting by Cas9 may lead to unexpected chromosomal translocations when targeting events occur on different chromosome locations.
O'Roak, B. J. et al. Multiplex targeted sequencing identifies recurrently mutated genes in autism spectrum disorders.
O'Roak, B.J. et al. Multiplex targeted sequencing identifies recurrently mutated genes in autism spectrum disorders.
Unlike ZFNs and TALENs, the CRISPR/Cas system is capable of multiplex gene targeting (Jao et al., 2013), which would allow for targeting of multiple tumor suppressors in a cancer model.
These results indicate that the RDB assay could be used to detect multiplex target genes of GMCs rapidly and inexpensively.
Therefore, we believe our method is ideally suited to multiplex target enrichment of samples with limited DNA resources, e.g. clinical samples.
In this study, we apply the SNaPshot multiplexed targeted sequencing platform in the analysis of brain tumors to interrogate 60 genetic loci that are frequently mutated in 15 cancer genes.
Collectively, this work shows that rationally engineered synthetic gene circuits are capable of treating multifactorial diseases in a synergistic manner by multiplexing the targeting efficacies of single therapeutics.
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