CRISPR-Cas9

Definition

CRISPR/Cas9 is a technique that allows for the highly specific and rapid modification of DNA in a genome, the complete set of genetic instructions in an organism.

This image depicts genome editing. It is adapted from a DNA illustration by Spooky Pooka. Credit: Wellcome Images.

Importance

The CRISPR/Cas 9 technique is one of a number of gene-editing tools. Many favour the CRISPR/Cas9 technique because of its high degree of flexibility and accuracy in cutting and pasting DNA. One of the reasons for its popularity is that it makes it possible to carry out genetic engineering on an unprecedented scale at a very low cost. How it differs from previous genetic engineering techniques is that it allows for the introduction or removal of more than one gene at a time. This makes it possible to manipulate many different genes in a cell line, plant or animal very quickly, reducing the process from taking a number of years to a matter of weeks. It is also different in that it is not species-specific, so can be used on organisms previously resistant to genetic engineering.

The technique is already being explored for a wide number of applications in fields ranging from agriculture through to human health. In agriculture it could help in the design of new grains, roots and fruits. Within the context of health it could pave the way to the development of new treatments for rare metabolic disorders and genetic diseases ranging from haemophilia through to Huntingdon's disease. It is also being utilised in the creation of transgenic animals to produce organs for transplants into human patients. The technology is also being investigated for gene therapy. Such therapy aims to insert normal genes into the cells of people who suffer from genetic disorders such as cystic fibrosis, haemophilia or Tay Sachs. Several start-up companies have been founded to exploit the technology commercially and large pharmaceutical companies are also exploring its use for drug discovery and development purposes.

Discovery

In 1987 a Japanese team of scientists at Osaka University noticed a strange pattern of DNA sequences in a gene belonging to Escherichia coli, a microbe that lives in the gut. It appeared that the gene had five short repeating segments of DNA separated by short non-repeating 'spacer' DNA sequences. All five repeating segments had identical sequences composed of 29 bases, the building blocks of DNA. By contrast each of the 'spacer' sequences had their own unique sequence, composed of 32 bases. Microbiologists had never seen such a pattern before. By the end of the 1990s, however, they had begun to discover, with the aid of new improvements to DNA sequencing, that this pattern was prevalent in many different microbe species.

So common was the pattern that it was given its own name: 'clustered regularly inter-spaced short palindromic repeats' or CRISPR for short. The term was coined by a team of Dutch scientists led by Rudd Jansen at Utrecht University, in 2002, who the same year noted that another set of sequences always accompanied the CRISPR sequence. This second set of sequences they dubbed 'Cas genes', an abbreviation for CRISPR-associated genes. The Cas genes appeared to code for enzymes that cut DNA. By 2005 three scientific teams had independently worked out that the 'spacer' sequences between the CRISP sequences shared similarities with the DNA of viruses and hypothesised that it could be a tool in the defence mechanism of bacteria.

Knowledge about how the CRISPR/Cas 9 system worked was opened up by some experiments conducted in 2007 by scientists at Danisco, a Danish food manufacturer later acquired by DuPont. The team infected a milk-fermenting microbe, Streptococcus thermophilius, with two virus strains. Many of these bacteria were killed by the viruses, but some survived and went on to produce offspring also resistant to the viruses. On further investigation it appeared that the microbes were inserting DNA fragments from the viruses into their 'spacer' sequences and that they lost resistance whenever the new 'spacer' sequences were cut out.

In 2008 Eugene Koonin and colleagues at the National Center for Biotechnology Information in Bethesda, Maryland, demonstrated for the first time how the CRISPR/Cas 9 mechanism worked. Whenever bacteria confront an invader, such as a virus, they copy and incorporate its DNA segments into their genome as 'spacers' between the short DNA repeats in CRISPR. The segments in the 'spacers' provide a template for the bacteria's RNA molecules to recognise any future DNA of an incoming virus and help guide the Cas 9 enzyme to cut it up so as to disable the virus.

Four years later, in 2012, a small team of scientists led by Jennifer Doudna, University California Berkeley, and Emmanuelle Charpentier, University of Umea, published a paper showing how to harness the natural CRISPR-Cas9 system as a tool to cut any DNA strand in a test tube. A year later, in January 2013, a number of researchers at different laboratories published papers within a few weeks of each other demonstrating how the CRISPR/Cas 9 system could be used to edit genomes in human cells. This included teams led by Doudna, Feng Zhang at MIT-Harvard Broad Institute, and George Church at Harvard Medical School.

A number of changes are now underway to improve the accuracy and efficiency of the CRISPR-Cas 9 technique. A key breakthrough has been the development of new Cas9 fusion proteins to act as base editors. The fusion proteins make it possible to convert cytosine to uracil without cutting DNA. Uracil is subsequently transformed into thymine through DNA replication or repair. The first base editors were generated in 2016 by Alexis Komor and colleagues in the laboratory of David Liu at Harvard University.

Application

The CRISPR/Cas 9 system was first exploited by Danisco in 2008. The company used it to improve the immunity of bacterial cultures against viruses and many food manufacturers now use the technology to produce cheese and yoghurt. Since then the technology has been used to delete, insert and modify DNA in human cells and other animal cells grown in petri dishes. Scientists are also using it to create transgenic animals such as mice, rats, zebrafish, pigs and primates. Between 2014 and 2015 scientists reported the successful use of CRISPR/Cas 9 in mice to eliminate muscular dystrophy and cure a rare liver disease, and to make human cells immune to HIV. It is also being investigated in conjunction with pluripotent stem cells to provide human organs from transgenic pigs. Such work is directed towards helping solve some of the shortage of human organs for transplant operations and overcome some of the side-effects caused by organ transplantation such as graft-versus host disease. The technology is also being investigated as a means to genetically engineer insects so as to wipe out insect-borne diseases such as malaria, transmitted by mosquitoes, and lyme disease, transmitted by ticks.

Issues

In April 2015 a Chinese group reported the first application of CRISPR/Cas9 to (non-viable) human embryos. This development, together with the decreasing costs of the technology have triggered a major bioethical debate about how far the technology should be used. The technology faces two major issues.

The first issue is a philosophical dilemma. It centres on the extent to which CRISPR/Cas9 should be used to alter 'germ-line' cells - eggs and sperm - which are responsible for passing genes on to the next generation. While it will take many more years before the technology will be viable to use to create designer babies, a public debate has already begun on this issue. So great is the fear that some scientists, including some who helped pioneer CRISPR/Cas9, have called for a moratorium on its use in germ-line cells.

The second issue is one of safety. One of the major problems is that the technology is still in its infancy and knowledge about the genome remains very limited. Many scientists caution that the technology still needs a lot of work to increase its accuracy and make sure that changes made in one part of the genome do not introduce changes elsewhere which could have unforeseen consequences. This is a particularly important issue when it comes to the use of the technology for applications directed towards human health. Another critical issue is that once an organism, such as a plant or insect, is modified they are difficult to distinguish from the wild-type and once released into the environment could endanger biodiversity.

Policy-makers are still debating about what limitations to put on the technology. In April 2015 the US National Institutes of Health issued a statement indicating that it will not fund any research that uses genome editing tools such as CRISPR in human embryos. Meanwhile, the UK's Human Fertilisation and Embryology Authority, under whose remit such research would fall, has indicated that the CRISPR/Cas9 technology can be used on human-animal hybrid embryos under 14 days old. Any researcher working in this area would need to first get a license from the Authority. Other leading UK research councils have indicated that they support the continued use of CRISPR/Cas9 and other genome editing tools in preclinical research.

As regulators debate what restrictions to enforce with CRISPR/Cas9, the technology has become the subject of a major patent dispute. The first application to patent the technology was filed by DuPont in March 2007 (WO/2007/025097). This covers the use of the technology to develop phage resistant bacterial strains for food production, feeds, cosmetics, personal care products and veterinary products. Since then three heavily financed start-up biotechnology companies and half a dozen universities have filed patents. Two major competing patent claims have been filed in the US. The first, filed on 25 May 2015, is grounded in the work led by Jennifer Doudna at the University of California, Berkeley, and Emmanuelle Charpentier, originally at the University of Vienna and now at the Helmholz Centre for Infectious Research in Germany. The application has 155 claims and covers numerous applications for a variety of cell types (US Patent Application No. PCT/US2013/032589). The second, was filed by MIT-Harvard Broad Institute on 12 December 2012 for the work of Feng Zhang which focused on the use of CRISPR/Cas9 for genome editing in eukaryotic cells. It was given fast-track status and was granted on 15 April 2014 (US Patent No. 8,697,359). In April 2015 Charpentier and the Universities of California and Vienna filed a challenge to the patent with the US Patent and Trademark Office. It will take several years for the patent dispute to be settled.

This profile was written by Lara Marks with generous input from Silvia Camporesi, Xiofan Zeng and Jonathan Lind.

CRISPR-Cas9: timeline of key events

Ishino Y, Shinagawa H, Makino K, Amemura M, Nakata A (December 1987). "Nucleotide sequence of the iap gene, responsible for alkaline phosphatase isozyme conversion in Escherichia coli, and identification of the gene product". Journal of Bacteriology 169 (12): 5429–5433; Takase, I, Ishino, F, Wachi, M, et al, 'Genes encoding two lipoproteins in the leuS-dacA region of the Escherichia coli chromosome', Journal of Bacteriology, 169/12 (1987): 5692-99.1987-12-01T00:00:00+0000Mojica F J, Diez-Villasenor C, Soria E, Juez G, 'Biological significance of a family of regularly spaced repeats in the genomes of Archaea, Bacteria and mitochondria', Molecular Microbiology 36/1 (April 2000): 244–6.2000-01-18T00:00:00+0000R Jansen, J D Embden, W Gaastra, L M, Schouls, 'Identification of genes that are associated with DNA repeats in prokaryotes', Molecular Microbiolology 43/6 (2002): 1565-75.2002-03-01T00:00:00+0000A Bolotin, B Quinquis, A Sorokin, S D Ehrlich, 'Clustered regularly interspaced short palindrome repeats (CRISPRs) have spacers of extrachromosomal origin', Microbiology, 151/Pt 8 (2005): 2551-6.2005-08-01T00:00:00+0000D H Haft, J Selengut, E F Mongodin, K E Nelson, 'A Guild of 45 CRISPR-Associated (Cas) Protein Families and Multiple CRISPR/Cas Subtypes Exist in Prokaryotic Genomes', PLoS Computational Biology, 1/6 (2005): e60. 2005-11-11T00:00:00+0000R Barrangou, C Fremaux, H Deveau, et al, 'CRISPR provides acquired resistance against viruses in prokaryotes', Science, 315/5819 (2007): 1709-12.2007-03-23T00:00:00+00002008-01-01T00:00:00+0000H Deveau, R Barrangou, J E Garneau, J Labonte et al, 'Phage Response to CRISPR-Encoded Resistance in Streptococcus thermophilus', Journal of Bacteriology, 190/4 (2008): 1390-1400.2008-02-01T00:00:00+0000S J J Brouns, M J Matthijs, M Lundgren, E R Westra et al, 'Small CRISPR RNAs guide antiviral defense in Prokaryotes', Science, 321/5891 (2008): 960-64.2008-08-15T00:00:00+0000J Carte, R Wang, H Li, R M and M P Terns, 'Cas6 is an endoribonuclease that generates guide RNAs for invader defense in prokaryotes', Genes Development, 22/24 (2008): 3489–96.2008-12-01T00:00:00+00002011-01-01T00:00:00+0000These are highly active Steptococcus thermophilus cultures.2012-04-02T00:00:00+0000The patent was submitted by Jennifer Doudna, at the University of California Berkeley, and Emmanuell Charpentier, at the Helmholtz Centre for Infection Research in Germany. The application was for a patent to cover the use of CRISPR-Cas9 for genome editing in vitro.2012-05-25T00:00:00+0000M Jinek, K Chylinski, I Fonfara, M Hauer, J A Doudna, E Charpentier, 'A programmable dual-RNA-guided DNA endonuclease in adaptive bacterial immunity', Science, 337/6096 (2012): 816-21.2012-08-17T00:00:00+0000Submitted by Feng Zhang at MIT. Covers method for using CRISPR-Cas9 for genome editing in eukaryotic cells. 2012-12-12T00:00:00+00002013-01-01T00:00:00+00002013-01-01T00:00:00+00002013-02-01T00:00:00+0000P Mali, L Yang, K M Esvelt, et al, 'RNA-guided human genome engineering via Cas9', Science, 339 (2013): 823-26.2013-02-15T00:00:00+00002013-03-01T00:00:00+00002013-04-01T00:00:00+00002013-08-01T00:00:00+00002013-08-01T00:00:00+0000Achieved by double-nicking with CRISP-Cas nickase mutant2013-08-01T00:00:00+0000Patent awarded to Feng Zhang of the Broad Institute and MIT.2014-03-01T00:00:00+0000W Feng, Y Dai, L Mou, D Cooper, D Shi, Z Cai, 'The potential of the combination of CRISPR/Cas9 and pluripotent stem cells to provide human organs from chimaeric pigs', International Journal of Molecular Science, 16/3 (2015): 6545-56.2015-03-23T00:00:00+0000E Lanphier, F Urnov et al, 'Don't edit the human germ line', Nature, 519 (2015): 410-11.2015-03-26T00:00:00+00002015-04-15T00:00:00+00002015-04-22T00:00:00+0000P Liang , Y Xu , X Zhang et al, 'CRISPR/Cas9-mediated gene editing in human tripronuclear zygotes', Protein and Cell, 6/5 (2015): 363-72. 2015-05-01T00:00:00+00002015-09-02T00:00:00+0000The group is made up of academic scientists, funders and regulators with an interest in embryo and stem cell research. It calls for the establishment of a roadmap for the management of the use of CRISPR in any human reproductive applications.2015-09-11T00:00:00+0000The group is made up of academic scientists, funders and regulators with an interest in embryo and stem cell research. It calls for the establishment of a roadmap for the management of the use of CRISPR in any human reproductive applications.2015-09-11T00:00:00+00002015-09-15T00:00:00+0000Permission sought from UK Human Fertilisation and Embryology Authority by scientists based at Francis Crick Institute in London. The research is to focus on embyos less than 2 weeks old and is designed to understand why some women lose their babies before term. 2015-09-18T00:00:00+0000B Zetsche, J Gootenberg, O Abudayyeh, I Slaymaker, et al, 'Cpf1 Is a Single RNA-Guided Endonuclease of a Class 2 CRISPR-Cas System', Cell (2015): 1-13.2015-09-25T00:00:00+0000Work presented by geneticist George Church of Harvard Medical School to US National Academy of Sciences.2015-10-05T00:00:00+0000Statement issued alongside its report 'Updating its Reflection on the Human Genome and Human Rights'. 2015-10-06T00:00:00+0000J E DiCarlo, A Chavez, S L Dietz, K M Esvelt, G M Church, 'Safeguarding CRISPR-Cas9 gene drives in yeast', Nature Biotechnology, doi:10.1038/nbt.3412.2015-11-16T00:00:00+0000V M Gantz, N Jasinskiene, O Tatarenkova, A Fazekas, V M Macias, E Bier, A A James, 'Highly efficient Cas9-mediated gene drive for population modification of the malaria vector mosquito Anopheles stephensi', PNAS Early edition (2015), doi/10.1073/pnas 15210771122015-11-23T00:00:00+0000Summit recommended the continuation of basic and clinical research of the applications gene editing in somatic cells, but stressed it would be 'irresponsible to proceed with any clinical use of germline editing' until the technology's safety and efficacy issues were better understood. 2015-12-01T00:00:00+0000Nelson, C E, Hakim, C H, Ousterout, D G, Thakore, P I, et al, 'In vivo genome editing improves muscle function in a mouse model of Duchenne muscular dystrophy', Science, DOI: 10.1126/science.aad51432015-12-31T00:00:00+0000Kleinstiver, B P, Pattanayak, V, Prew, M S, Tsai, S Q, Nguyen, N T, Zheng, Z, Joung, K, 'High-fidelity CRISPR–Cas9 nucleases with no detectable genome-wide off-target effects', Nature, doi:10.1038.2016-01-06T00:00:00+0000The UK Human Fertilisation and Embryology Authority give the go ahead to a team at the Crick institute, led by Kathy Nikan, to edit human embyos to undestand the genetic cause of miscarriage. 2016-02-01T00:00:00+0000A.C. Komor, Y.B. Kim, M.S. Packer, J.A. Zuris, D.R. Liu, 'Programmable editing of a target base in genomic DNA without double-stranded DNA cleavage', Nature, 533/7603 (2016), 420–24.2016-05-16T00:00:00+0000Work led by Carl June at University of Pennsylvannia to treat patients with multiple myeloma, melanoma and sarcoma.2016-06-21T00:00:00+0000NAS and NAM, Human Genome Editing: Science, Ethics and Governance, February 20172017-02-24T00:00:00+0000J. S. Gootenberg, O.O. Abudayyeh, J. W. Lee, et al, 'Nucleic acid detection with CRISPR-Cas13a/C2c2', Science, 13 April 2017, eaam9321, DOI: 10.1126/science.aam9321 2017-04-13T00:00:00+0000The experiment was carried out by scientists at Sichuan University, Temple University and the University of Pittsburgh. It was published in C. Yin, et al, 'In Vivo Excision of HIV-1 Provirus by saCas9 and Multiplex Single-Guide RNAs in Animal Models', Molecular Therapy, 25/5 (2017), 1168-86.2017-05-13T00:00:00+0000M. Hong, N. Marti-Gutierrez, S-W Park, et al, 'Correction of a pathogenic gene mutation in human embryos', Nature, doi:10.1038/nature233052017-08-02T00:00:00+0000UK scientists modified 41 embryos shortly after fertilisation. N.M.E. Fogarty et al, 'Genome editing reveals a role for OCT4 in human embryogenesis', Nature, doi:10.1038/nature240332017-09-20T00:00:00+0000P. Liang, et al, 'Correction of beta-thalassemia mutant by base editor in human embryos', Protein and Cell (2017), doi.org/10.1007/s13238-017-0475-6.2017-09-23T00:00:00+0000
Date Event People Places
December 1987Discovery of CRISPR mechanismAmemura, Ishino, Makino, Nakata, Shinagawa, Takase, WachiOsaka University
18 Jan 2000More clustered repeats of DNA identified in other bacteria and archaea, termed Short Regularly Spaced Repeats (SRSR)Mojica, Diez-Villasenor, Soria, Juez University of Alicante, University Miguel Hernandez
March 2002Dutch scientists coin the term CRISPR-Cas9. Strands for clustered regularly interspaced short palindrome repeats in a DNA sequence and its associated genes.Jansen, Embden, Gaastra, SchoulsUtrecht University
August 2005French scientists suggest CRISPR spacer sequences provide cell immunity against phage infection and are a means of degrading DNABolotin, Quinquis, Sorokin, EhrlichInstitut National de la Recherche Agronomique
November 2005American researchers identified new familes of Cas genes which appeared to help in protecting bacteria against invading virusesHaft, Selengut, Mongodin, NelsonThe Institute for Genomic Research
March 2007Experiments conducted by microbiologists based at the food manufacturing company Danisco, demonstrate for the first time the role of CRISPR together with Cas9 genes in protecting bacteria against virusesBarrangou, Horvath, Fremaux, Deveau, Danisco USA Inc
2008DNA, not RNA, demonstrated to be the molecular target of most CRISPR-Cas systems 
February 2008Scientists coin the term 'protospacer' to denote viral sequence that corresponds to a 'spacer' in the CRISPR-Cas9 system 
August 2008Scientists characterised the RNA processing pathway in CRISPR systemWageningen University, University of Sheffield, National Institutes of Health
December 2008Scientists published the RNA gene silencing pathway involved in the CRISPR-Cas mechanism University of Georgia, Florida State University
2011Classification of the CRISPR-Cas system is proposed 
April 2012DuPont commercialises the first bacterial cultures based on the CRISPR-Cas 9 technology for the production of pizza cheeseDupont
May 2012First patent application submitted for CRISPR-Cas 9 technologyDoudna, CharpentierUniversity of California Berkeley, University of Vienna
August 2012A group of scientists based at Howard Hughes Medical Institute published a radically new gene editing method that harnessed the CRISPR-Cas9 system Jinek, Chylinski, Fonfara, Hauer, Doudna, CharpentierUniversity of California Berkeley
December 2012Fast track application for CRISPR-Cas 9 technology submitted to US patent office. ZhangBroad Institute, Massachusetts Institute of Technology
January 2013CRISPR-Cas is used in human genome editing 
January 2013CRISPR-Cas is used to edit the genome of a zebrafish 
February 2013CRISPR-Cas is used to programme repression and activation of gene transcription  
February 2013Scientists from Harvard Medical School announce the successful genetic engineering of human cells using a CRISPR-Cas 9 techniqueMali, Yang, Esvelt, Aach, Guell, DiCarlo, Norville, ChurchHarvard University
March 2013CRISPR-Cas is used in genome editing of Saccharomyces cerevisiae, a yeast species used in wine making, baking and brewing 
April 2013Scientists show that CRISPR-Cas mediated gene regulation contributes to the regulation of endogenous bacterial genesWeiss 
August 2013CRISPR-Cas used to engineer a rat's genome 
August 2013CRISPR-Cas used to engineer plant genomes including rice, wheat, Arabidopsis, tobacco and Sorghum 
August 2013Improvements made to the specificity of CRISPR-Cas system 
March 2014First patent awarded for CRISPR-Cas9 technology to be used for a wide range of applicationsZhangBroad Institute, Massachusetts Institute of Technology
March 2015Scientists suggest the use of CRISPR/Cas9 in conjunction with stem cells to provide human organs from transgenic pigsFeng, Dai, Mou, Cooper, Shi, Cai Shenzhen University, University of Pittsburgh Medical Center, Guangxi University
26 Mar 2015US scientists call for a voluntary worldwide moratorium on the use of genome editing tools to modify human reproductive cellsLamphier, Urnov 
April 2015National Institutes of Health declares it will not fund any use of genome editing technologies in human embryos 
22 Apr 2015UK Nuffield Council on Bioethics launched a new working group to look into institutional, national and international policies and provisions relevant to genome editing 
May 2015Chinese scientists, led by Junjiu Huang, publish first experiments modifying the genome of human embryos with CRISPR-Cas9 technique, prompting bioethical debate over use of the technology Huang, Liang, Xu, ZhangSun Yat-sen University
September 2015UK Medical Research Council and other leading UK research councils declared support for using CRISPR-Cas9 and other genome editing techniques in preclinical research 
11 Sep 2015Hinxton Group issues a statement indicating that most of the ethical and moral questions raised about CRISPR and gene editing have been debated before 
11 Sep 2015Hinxton Group issues a statement indicating that most of the ethical and moral questions raised about CRISPR and gene editing have been debated before 
15 Sep 2015UK Nuffield Council on Bioethics held its first workshop to identify and define ethical questions relating to developments in genome editing research 
18 Sep 2015UK scientists seek license to genetically modify human embryos to study the role played by genes in the first few days of human fertilisationNaikanCrick Institute
25 Sep 2015New protein, Cpf1, found which could simplify genome editing. Zhang, Zetsche, Gootenberg, Abudayyeh, SlaymakerBroad Institute, Massachusetts Institute of Technology
5 Oct 2015US Scientists report use of CRISPR to modify 60 genes in pig embryos in first step to create pig organs suitable for use in humansChurchHarvard University
6 Oct 2015UNESCO’s International Bioethic Committee called for ban on genetic editing of human germline  
16 Nov 2015US Scientists publish a technique for overwriting changes made by CRISPR/Cas 9DiCarlo, Chavez, Dietz, Esvelt, ChurchHarvard University, Swiss Federal Institute of Technology in Zurich
23 Nov 2015US scientists report successful use of CRISPR/Cas9 to genetically modify mosquitoes to make them poor hosts for malaria parasiteGantz, Jasinskiene, Tatarenkova, Fazekas, Macias, Bier, JamesUniversity California San Diego, University of California Irvine
1 Dec 2015 - 3 Dec 2015 International Summit on Human Gene Editing met to discuss the scientific, medical, ethical, and governance issues associated with recent advances in human gene-editing researchBaltimore, Doudna, Church, ZhangUS National Academies of Science, Engineering and Medicine, US National Academy of Medicine, Chinese Academy of Sciences, Royal Society
31 Dec 2015CRISPR successfully used to improve muscle function in mouse model of Duchenne muscular dystrophy, opening way to use CRISPR to correct genetic mutatiuons in affected tissues of sick patientsNelson, Gersbach, Hakim, Ousterout, ThakoreDuke University, University of Missouri, University of North Carolina, Massachusetts Institute of Technology, Harvard University
6 Jan 2016US scientists published improved version of CRISPR/Cas 9 with less risk of off-target DNA breaksKleinstiver, Pattanayak, Prew, Tsai, Nguyen, Zheng, JoungHarvard University
1 Feb 2016UK scientists authorised to genetically modify human embryos using CRISPR-Cas 9NiakanCrick Institute
16 May 2016US scientists publish new base editing technique offering means to alter genome without needing to cleave double-stranded DNA or for a donor DNA templateKomor, Kim, Packer, Zuris, LiuHarvard University
21 Jun 2016US NIH Recombinant DNA Advisory Committee approved first study in 15 patients using CRISPR/Cas 9 to genetically modify immune cells to attack cancerJuneUniversity of Pennsylvania
February 2017US National Academies of Science and Medicine issue report saying possible to proceed with CRISPR in germ-line experiments but urge caution 
13 Apr 2017CRISPR shown to be sensitive diagnostic tool for detecting single target of DNA or RNA moleculeAbudayyeh, Bhattacharyya, Collins, Daringe, Donghia, Dy, Essletzbichler, Freije, Hung, Joung, Koonin, Lee, Livny, Myhrvold, Regev, Sabeti, Gootenberg, Verdine, ZhangBroad Institute, Massachusetts Institute of Technology, Harvard University, Howard Hughes Medical Institute
13 May 2017Research published demonstrating how CRISPR-CAS9 can be used to eliminate HIV in infected mice. Yin, Zhang, Qu, Chang, Putatunda, Xiao, Li, Zhao, Dhai, Qin, Mo, Young, Khalili, HuTemple University, University of Pittsburgh, Sichuan University
2 Aug 2017Research published demonstrating possibility of editing gene defect in pre-implanted human embryos for preventing inherited heart diseaseHong, Marti-Gutierrez, Park, Mitalipov, Kaul, Kim, Amato, BelmonteOregon Health & Science University, Salk Institute, Center for Genome Engineering, Seoul National University, China National GeneBank,
September 2017DNA of human embryos edited using CRISPR-Cas9 to study cause of infertilityFogarty, McCarthy, Snijders, Powell, Kubikova, Blakeley, Lea, Elder, Wamaitha, Kim, Maciulyte, Kleinjung, Kim, Wells, Vallier, Bertero, Turner, NiakanFrancis Crick Instiitute, Cambridge University, Oxford University, Seoul National University
23 Sep 2017Chinese researchers report correction of gene linked to beta thalassaemia, inherited blood disorder, in human embryos using base editing techniqueLiang, Ching, Sun, Xie, Xu, Zhang, Xhiong, Ma, Liu, Wang, Fang, Songyang, Zhou, HuangSun Yat-sen University, Baylor College of Medicine

Dec 1987

Discovery of CRISPR mechanism

18 Jan 2000

More clustered repeats of DNA identified in other bacteria and archaea, termed Short Regularly Spaced Repeats (SRSR)

Mar 2002

Dutch scientists coin the term CRISPR-Cas9. Strands for clustered regularly interspaced short palindrome repeats in a DNA sequence and its associated genes.

Aug 2005

French scientists suggest CRISPR spacer sequences provide cell immunity against phage infection and are a means of degrading DNA

Nov 2005

American researchers identified new familes of Cas genes which appeared to help in protecting bacteria against invading viruses

Mar 2007

Experiments conducted by microbiologists based at the food manufacturing company Danisco, demonstrate for the first time the role of CRISPR together with Cas9 genes in protecting bacteria against viruses

2008

DNA, not RNA, demonstrated to be the molecular target of most CRISPR-Cas systems

Feb 2008

Scientists coin the term 'protospacer' to denote viral sequence that corresponds to a 'spacer' in the CRISPR-Cas9 system

Aug 2008

Scientists characterised the RNA processing pathway in CRISPR system

Dec 2008

Scientists published the RNA gene silencing pathway involved in the CRISPR-Cas mechanism

2011

Classification of the CRISPR-Cas system is proposed

Apr 2012

DuPont commercialises the first bacterial cultures based on the CRISPR-Cas 9 technology for the production of pizza cheese

May 2012

First patent application submitted for CRISPR-Cas 9 technology

Aug 2012

A group of scientists based at Howard Hughes Medical Institute published a radically new gene editing method that harnessed the CRISPR-Cas9 system

Dec 2012

Fast track application for CRISPR-Cas 9 technology submitted to US patent office.

Jan 2013

CRISPR-Cas is used in human genome editing

Jan 2013

CRISPR-Cas is used to edit the genome of a zebrafish

Feb 2013

CRISPR-Cas is used to programme repression and activation of gene transcription

Feb 2013

Scientists from Harvard Medical School announce the successful genetic engineering of human cells using a CRISPR-Cas 9 technique

Mar 2013

CRISPR-Cas is used in genome editing of Saccharomyces cerevisiae, a yeast species used in wine making, baking and brewing

Apr 2013

Scientists show that CRISPR-Cas mediated gene regulation contributes to the regulation of endogenous bacterial genes

Aug 2013

CRISPR-Cas used to engineer a rat's genome

Aug 2013

CRISPR-Cas used to engineer plant genomes including rice, wheat, Arabidopsis, tobacco and Sorghum

Aug 2013

Improvements made to the specificity of CRISPR-Cas system

Mar 2014

First patent awarded for CRISPR-Cas9 technology to be used for a wide range of applications

Mar 2015

Scientists suggest the use of CRISPR/Cas9 in conjunction with stem cells to provide human organs from transgenic pigs

26 Mar 2015

US scientists call for a voluntary worldwide moratorium on the use of genome editing tools to modify human reproductive cells

Apr 2015

National Institutes of Health declares it will not fund any use of genome editing technologies in human embryos

22 Apr 2015

UK Nuffield Council on Bioethics launched a new working group to look into institutional, national and international policies and provisions relevant to genome editing

May 2015

Chinese scientists, led by Junjiu Huang, publish first experiments modifying the genome of human embryos with CRISPR-Cas9 technique, prompting bioethical debate over use of the technology

Sep 2015

UK Medical Research Council and other leading UK research councils declared support for using CRISPR-Cas9 and other genome editing techniques in preclinical research

11 Sep 2015

Hinxton Group issues a statement indicating that most of the ethical and moral questions raised about CRISPR and gene editing have been debated before

11 Sep 2015

Hinxton Group issues a statement indicating that most of the ethical and moral questions raised about CRISPR and gene editing have been debated before

15 Sep 2015

UK Nuffield Council on Bioethics held its first workshop to identify and define ethical questions relating to developments in genome editing research

18 Sep 2015

UK scientists seek license to genetically modify human embryos to study the role played by genes in the first few days of human fertilisation

25 Sep 2015

New protein, Cpf1, found which could simplify genome editing.

5 Oct 2015

US Scientists report use of CRISPR to modify 60 genes in pig embryos in first step to create pig organs suitable for use in humans

6 Oct 2015

UNESCO’s International Bioethic Committee called for ban on genetic editing of human germline

16 Nov 2015

US Scientists publish a technique for overwriting changes made by CRISPR/Cas 9

23 Nov 2015

US scientists report successful use of CRISPR/Cas9 to genetically modify mosquitoes to make them poor hosts for malaria parasite

1 Dec 2015 - 3 Dec 2015

International Summit on Human Gene Editing met to discuss the scientific, medical, ethical, and governance issues associated with recent advances in human gene-editing research

31 Dec 2015

CRISPR successfully used to improve muscle function in mouse model of Duchenne muscular dystrophy, opening way to use CRISPR to correct genetic mutatiuons in affected tissues of sick patients

6 Jan 2016

US scientists published improved version of CRISPR/Cas 9 with less risk of off-target DNA breaks

1 Feb 2016

UK scientists authorised to genetically modify human embryos using CRISPR-Cas 9

16 May 2016

US scientists publish new base editing technique offering means to alter genome without needing to cleave double-stranded DNA or for a donor DNA template

21 Jun 2016

US NIH Recombinant DNA Advisory Committee approved first study in 15 patients using CRISPR/Cas 9 to genetically modify immune cells to attack cancer

Feb 2017

US National Academies of Science and Medicine issue report saying possible to proceed with CRISPR in germ-line experiments but urge caution

13 Apr 2017

CRISPR shown to be sensitive diagnostic tool for detecting single target of DNA or RNA molecule

13 May 2017

Research published demonstrating how CRISPR-CAS9 can be used to eliminate HIV in infected mice.

2 Aug 2017

Research published demonstrating possibility of editing gene defect in pre-implanted human embryos for preventing inherited heart disease

Sep 2017

DNA of human embryos edited using CRISPR-Cas9 to study cause of infertility

23 Sep 2017

Chinese researchers report correction of gene linked to beta thalassaemia, inherited blood disorder, in human embryos using base editing technique