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CRISPR is the abbreviation of <u>'''C'''</u>lustered <u>'''R'''</u>egularly <u>'''I'''</u>nterspaced <u>'''S'''</u>hort [[Palindromic sequence|<u>'''P'''</u>alindromic <u>'''R'''</u>epeats]], a mediated defense mechanism adopted by prokaryotes.The widely applied CRISPR technology is the Crispr/Cas system for gene sequence editions by transferring [[Cas9 protein|Cas9 protein]] family encoded gene into cells and potentially simplifies the procedure of cutting [[DNA|DNA]] at a desired location<ref>Ledford H (3 June 2015). "CRISPR, the disruptor". News Feature. Nature 522 (7554).</ref>. Cas9 was first used for targeted genome editing in prokaryotes by Prof. Emmanuelle Charpentier &amp; Prof. Jennifer Doudna (2012) and later adapted for use in eukaroytes by Prof. Feng Zhang (2013). 

There are three CRISPR systems, one of which has been adapted for eukaryotes. This is the [[Type II CRISPR|Type II CRISPR]] system, which involves Cas9 protein, and it consists of three stages summarised briefly below: 

#Acquisition, where the invading [[DNA|DNA]] is recognised by Cas 1 and 2, and a protospacer is cleaved. The protospace then ligates to the repeat directly opposite the leader sequence, and single-stranded extension repairs the CRISPR array and duplicates the repeat (forming pre-crRNA). A copy of the invader's DNA is now incorporated into CRISPR's locus. 
#crRNA processing, where tracrRNA hybridizes to the repeat regions of the pre-crRNA, and with the help of RNAse III, mature crRNAs with individual spacer sequences are created. Note: there will be a secondary trimming performed at the 5'-end to produce mature crRNAs. 
#Interference, where the mature crRNA:tracrRNA complex directs cas9 to the target DNA via base pairing between the spacer on the crRNA and the protospacer on the target DNA next to the PAM (Protospacer Adjacent Motif, is the sequence 5'-NGG-3' where "N" is any base). Finally, Cas9 mediates cleavage of target DNA to create a double-stranded break within the protospacer.

The DNA repairs this [[Double stranded break|double stranded break]] (DSB) by either [[Non-homologous end joining|non-homologous end joining]] (NHEJ) or [[Homology directed repair|homology directed repair]] (HDR). CRISPR uses NHEJ to knockout a gene from an organism, whereas HDR is used to insert a gene into an organism's genome. More differences between the two are highlighted [http://emendobio.com/technology/dna-repair/ here]<ref name="genome editing">Bollen Y, Post J, Koo B, Snippert H: How to create state-of-the-art genetic model systems: strategies for optimal CRISPR-mediated genome editing. Nucleic Acids Research (2018) 46:6435-6454.</ref>&nbsp; 

The CRISPR technology is now being used in genome editing, and in particular to fight disease. For example, specific gRNA molecules are being specifically designed alongside Cas9 to target [[Antibiotic resistance|antibiotic resistance]] genes in prokaryotes. CRISPR technology is also being adapted to treat genetic diseases such as Duchenne Muscular Dystrophy (DMD), which causes a lack in dystrophin which helps to protect muscles from injury, through in vivo genome editing. Caused by a mutation inactivating the dystrophin gene on the X chromosome, the injection of viral vectors coding for CRISPR/Cas9 is seeing early success in treating mice<ref>Nelson CE, Hakin CH, Ousterout DG, Thakore PI, Moreb EB et al. In vivo genome editing improves muscle function in mouse model of Duchenne muscular dystrophy. Science 2016; 351(6271)pp403-407: http://science.sciencemag.org/content/351/6271/403</ref>. Alongside this, labs are trying to modify mosquitos to release into the wild in order to eradicate [[Malaria|malaria]], however, there are some ethical concerns stipulating this e.g. "designer babies" using genome editing. As a pioneer of the CRISPR/Cas9 technology, Prof. Jennifer Doudna responsibly called for a "global pause" in any clinical application of the CRISPR technology in human embryos to give time to consider the implications of doing so, just as scientists did in the 70s to consider the use of molecular cloning. However, she did say during a TED talks on CRISPR in 2015 that even though genome-engineered humans aren't with us yet, "this is no longer science fiction" and that "in the end, this technology will be used for human genome engineering". 

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