maybe theses articles will be useful

http://www.nature.com/nbt/journal/v32/n4/full/nbt.2842.html

http://www.addgene.org/CRISPR/?gclid=CNyZ_ceXtMkCFQyAkQodEB0AFw

http://www.cell.com/nucleus-CRISPR

Yes and no.  NHEJ should work fine in non-cycling cells so INDEL production using CRISPR would be pretty effective.

Homologous recombination, which is needed for homology-directed repair or precision editing of genes does not work in non-dividing cells because 53BP1 inhibits DNA end resection and recruitment of BRCA1 to breaks.   In addition the KEAP1/CUL3 E3 ligase complex inhibits the interaction between BRCA1 and PALB2 through ubiquitinylation of PALB2.  Without manipulating these pathways in the G1 (or post-mitotic) cells (specifically Keap1/CUL3 inhibition, knockout of 53BP1 and some means of enhancing resection, e.g. CtIP mutants) you cannot do CRISPR targeting using homologous recombination.  

see: Orthwein et al., 2015 Nature

Then once you have homologous recombination working again, you can enhance HR and CRISPR targeting using a RAD51 enhancer such as RS-1. See: Pinder et al., 2015 NAR

http://www.nature.com/nature/journal/vaop/ncurrent/full/nature16142.html

http://nar.oxfordjournals.org/content/43/19/9379.long

Dear Reader,

Thank you for your inquiry regarding the application of genome editing technologies, specifically CRISPR/Cas9, in non-cycling (quiescent) cells. This is a pertinent question, especially given the expanding scope of CRISPR/Cas9 applications in genetics, biomedicine, and therapeutic interventions.

CRISPR/Cas9 has revolutionized the field of genetic engineering by providing a highly precise, efficient, and versatile tool for editing the genome of various organisms. The system works by utilizing a guide RNA (gRNA) to direct the Cas9 nuclease to a specific DNA sequence, where it induces a double-strand break. The cell then repairs this break, allowing for the introduction of mutations or the insertion of new genetic material.

The effectiveness of CRISPR/Cas9 in non-cycling cells, which are cells in a quiescent state not actively dividing, is a subject of keen interest. Historically, the efficiency of many genome editing technologies, including CRISPR/Cas9, was thought to be higher in proliferating cells. This belief stems from the mechanisms involved in DNA repair pathways—specifically, the preference for homology-directed repair (HDR) in dividing cells, which is often used in precise genome editing strategies.

However, recent studies have demonstrated that CRISPR/Cas9 can indeed be effective in non-cycling cells. The key lies in the cell's ability to engage non-homologous end joining (NHEJ) or microhomology-mediated end joining (MMEJ), alternative DNA repair pathways that are active in both dividing and non-dividing cells. While these pathways are generally considered less precise than HDR, advancements in CRISPR/Cas9 delivery methods, gRNA design, and Cas9 variants have improved the precision and efficiency of edits in non-cycling cells.

Applications in non-cycling cells are particularly relevant for therapeutic interventions targeting tissues composed largely of quiescent cells, such as neurons, muscle cells, and hepatocytes. Successful genome editing in these cells could open new avenues for treating genetic disorders, neurodegenerative diseases, and muscular dystrophies, among others.

In summary, while the efficiency and mechanisms of genome editing with CRISPR/Cas9 may vary between cycling and non-cycling cells, ongoing research and technological advancements have made it increasingly feasible to edit the genome of non-cycling cells. This expands the potential applications of CRISPR/Cas9 far beyond traditional models, promising significant advancements in genetic research and therapeutic development.

Best regards,

With this protocol list, we might find more ways to solve this problem.