CRISPR-A gene editing tool
Clustered Regularly Interspaced Short Palindromic Repeats or CRISPR is a genome editing system that acts as an immune system against invading viruses and plasmids in prokaryotic organisms such as bacteria and archaea. The CRISPR system involves two components: a single guide RNA (sgRNA) with complementarity to any sequence in the genome, and the Cas9 (CRISPR-associated) endonuclease, which associates with the sgRNA at the genomic target sequence. Francisco Mojica first identified it in Escherichia coli bacteria (present in the human body) in 1993. While sequencing the DNA of a halophilic microorganism of the archaea group, he observed regularly spaced repeats which would later be called CRISPR. Precision targeting and the creation of double-strand breaks for gene manipulation are the two most vital requirements in genome editing, and CRISPR fulfils both, making it a revolutionary discovery.
In 2020, Jennifer Doudna and Emmanuelle Charpentier won a Nobel Prize in Chemistry for their landmark paper published in 2012, which popularised CRISPR. They had identified a key component in the CRISPR system other than the Cas protein, an RNA molecule involved in recognizing phage sequences. The paper proved that although CRISPR is not the first gene editing tool, it could revolutionize the field with its ease of functionality, quick results, and minimal expenses. The CRISPR gene-editing tool has numerous medical applications, such as the treatment of HIV infection, resistance against malaria, a possible cure for cancer, the potential eradication of genetic disorders, and many more.
In 2013, the CRISPR tool was further developed and applied to modify genes in several model organisms such as Drosophila, Caenorhabditis elegans, and zebrafish. Simply injecting Cas9 mRNA and gRNA into early embryos resulted in efficient genome editing. β-thalassemia, one of the most common genetic diseases worldwide, is caused by mutations in the human haemoglobin beta (HBB) gene. In 2014 researchers successfully edited the first non-viable human embryo using CRISPR to correct the mutation that leads to beta-thalassemia. Also, in 2019 researchers figured out CRISPR could also correct mutations in the X-linked dystrophin gene that cause Duchenne muscular dystrophy (DMD). DMD is the most common lethal monogenic disorder, primarily affecting boys owing to their single X chromosome. The incidence of the disease is estimated at 1:5,000 boys worldwide. Approximately two-thirds of DMD mutations are inherited by the sons of mothers who are unknowing carriers of dystrophin mutations.
CRISPR-Cas modules are adaptive antivirus immunity systems based on the self-nonself discrimination principle. This system is divided into two classes. Class I and Class II. Class I is divided into Types I, III, and IV. These are further divided into subtypes. Class II is divided into Type II, Type V and Type VI. Even with such variety, due to the simplicity, high efficiency, and multiplexing capability of the type II CRISPR/Cas system, it has been adopted as the genome editing technology of choice. The type II system utilizes a single Cas9 protein nuclease sufficient to cleave the target DNA specified by crRNA.