vendredi 4 septembre 2015

More on CRISPR

This recent review in Cell by Zhang exemplifies the wide spectrum of CRISPR use in DNA editing.
Applications of Cas9 as a Genome Engineering Platform(A) The Cas9 nuclease ...
Figure 6. 
Applications of Cas9 as a Genome Engineering Platform
(A) The Cas9 nuclease cleaves DNA via its RuvC and HNH nuclease domains, each of which nicks a DNA strand to generate blunt-end DSBs. Either catalytic domain can be inactivated to generate nickase mutants that cause single-strand DNA breaks.
(B) Two Cas9 nickase complexes with appropriately spaced target sites can mimic targeted DSBs via cooperative nicks, doubling the length of target recognition without sacrificing cleavage efficiency.
(C) Expression plasmids encoding the Cas9 gene and a short sgRNA cassette driven by the U6 RNA polymerase III promoter can be directly transfected into cell lines of interest.
(D) Purified Cas9 protein and in vitro transcribed sgRNA can be microinjected into fertilized zygotes for rapid generation of transgenic animal models.
(E) For somatic genetic modification, high-titer viral vectors encoding CRISPR reagents can be transduced into tissues or cells of interest.
(F) Genome-scale functional screening can be facilitated by mass synthesis and delivery of guide RNA libraries.

Applications of Genome EngineeringGenetic and epigenetic control of cells with ...

Figure 1.
Applications of Genome Engineering
Genetic and epigenetic control of cells with genome engineering technologies is enabling a broad range of applications from basic biology to biotechnology and medicine. (Clockwise from top) Causal genetic mutations or epigenetic variants associated with altered biological function or disease phenotypes can now be rapidly and efficiently recapitulated in animal or cellular models (Animal models, Genetic variation). Manipulating biological circuits could also facilitate the generation of useful synthetic materials, such as algae-derived, silica-based diatoms for oral drug delivery (Materials). Additionally, precise genetic engineering of important agricultural crops could confer resistance to environmental deprivation or pathogenic infection, improving food security while avoiding the introduction of foreign DNA (Food). Sustainable and cost-effective biofuels are attractive sources for renewable energy, which could be achieved by creating efficient metabolic pathways for ethanol production in algae or corn (Fuel). Direct in vivo correction of genetic or epigenetic defects in somatic tissue would be permanent genetic solutions that address the root cause of genetically encoded disorders (Gene surgery). Finally, engineering cells to optimize high yield generation of drug precursors in bacterial factories could significantly reduce the cost and accessibility of useful therapeutics (Drug development).

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