Using CRISPR/Cas9 to Edit the Gene of the Primary Cell
Using CRISPR/Cas9 system for gene editing of embryos is effective to generate organisms with targeted genome modification.
- (1888PressRelease) February 07, 2016 - Shirley, NY - In 2015, Cell Reports published a study that researchers from the University of Texas Southwestern Medical Center, the University of Chicago and the University of Utah, edit gene of rat spermatogonial cells using CRISPR/Cas9 technology, providing a platform for producing targeted germline mutation and the spermatogenesis in rats.
CRISPR/cas9 RNA guided DNA enzyme technology is widely used to generate targeted gene mutations in different types of cells and organisms to study their biological processes. In mammals, we use CRISPR / cas9 to edit gene. In rodents, pigs, goats and monkeys have a high rate of donor - embryo for offspring, and they have a target gene mutation. Mice injected with gRNAs and cas9 are able to produce variation less than a month.
CRISPR/cas9 also can effectively promote the targeted alleles mosaic in animals, suggesting that totipotent zygote undergoes preimplantation development and early cleavage stage embryos occurred independent gene editing activities. CRISPR/cas9 is usually delivered to the mammalian fertilized egg at the age of E0.5 days. For example, during E6 to E6.5, mouse germ cells differentiate into a small portion of the outer germ layer cells (10 to 20 proximal germ cells).
This explains why, in the first generation of mutant animals, the CRISPR / cas9 target allele is embedded, and why the target allele in the body tissue is different from that of the germ cell. The heterogeneity of the target alleles in the mosaic animals must be produced by the distant cross to produce a pure, non mosaic mutant gene. By experimenting outcrossing allele mosaic to select new mutations, it needs to spend a few months in rodents. In some species, due to a longer life cycle or low fecundity, it still needs a longer period of time.
When the host embryo is reconstructed with CRISPR transgenic pluripotent stem cells, it also generates a target allele. As early embryos, CRISPR / cas9 may significantly modify multiple target genes in stem cell clones. In addition to this variation, wild-type blastocyst constructed by multi donor cell can further produce "mosaic" animals and their organ systems and reproductive cells are developed by donor and acceptor embryo cell mixture.
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