Editing Germline mutation (section)

=== CRISPR/Cas9 editing ===
[[File:DNA Repair after CRISPR-Cas9 cut.svg|thumb|254x254px|The CRISPR editing system is able to target specific DNA sequences and, using a donor DNA template, can repair mutations within this gene.]]
This editing system induces a double stranded break in the DNA, using a guide RNA and effector protein Cas9 to break the DNA backbones at specific target sequences.<ref name=":6" /> This system has shown a higher specificity than TALENs or ZFNs due to the Cas9 protein containing homologous (complementary) sequences to the sections of DNA surrounding the site to be cleaved.<ref name=":6">{{cite journal | vauthors = Sander JD, Joung JK | title = CRISPR-Cas systems for editing, regulating and targeting genomes | journal = Nature Biotechnology | volume = 32 | issue = 4 | pages = 347–55 | date = April 2014 | pmid = 24584096 | doi = 10.1038/nbt.2842 | pmc=4022601}}</ref>&nbsp;This broken strand can be repaired in 2 main ways: homologous directed repair (HDR) if a DNA strand is present to be used as a template (either homologous or donor), and if one is not, then the sequence will undergo [[non-homologous end joining]] (NHEJ).<ref name=":6" /> NHEJ often results in insertions or deletions within the gene of interest, due to the processing of the blunt strand ends, and is a way to study gene knockouts in a lab setting.<ref name=":5">{{cite journal | vauthors = Shalem O, Sanjana NE, Hartenian E, Shi X, Scott DA, Mikkelson T, Heckl D, Ebert BL, Root DE, Doench JG, Zhang F | title = Genome-scale CRISPR-Cas9 knockout screening in human cells | journal = Science | volume = 343 | issue = 6166 | pages = 84–87 | date = January 2014 | pmid = 24336571 | doi = 10.1126/science.1247005 | pmc=4089965| bibcode = 2014Sci...343...84S }}</ref> This method can be used to repair a point mutation by using the [[Chromatid|sister chromosome]] as a template, or by providing a double stranded DNA template with the [[CRISPR]]/Cas9 machinery to be used as the repair template.<ref name=":6" />

This method has been used in both human and animal models (''[[Drosophila]]'', ''[[House mouse|Mus musculus]]'', ''and [[Arabidopsis]]''), and current research is being focused on making this system more specific to minimize off-target cleavage sites.<ref>{{cite journal | vauthors = Smith C, Gore A, Yan W, Abalde-Atristain L, Li Z, He C, Wang Y, Brodsky RA, Zhang K, Cheng L, Ye Z | title = Whole-genome sequencing analysis reveals high specificity of CRISPR/Cas9 and TALEN-based genome editing in human iPSCs | language = en | journal = Cell Stem Cell | volume = 15 | issue = 1 | pages = 12–3 | date = July 2014 | pmid = 24996165 | doi = 10.1016/j.stem.2014.06.011 | pmc=4338993}}</ref>