Decoding the Mechanism- How CRISPR Accurately Targets the Gene of Interest
How Does CRISPR Target the Gene of Interest?
CRISPR (Clustered Regularly Interspaced Short Palindromic Repeats) technology has revolutionized the field of genetic engineering by providing a precise and efficient method for editing genes. One of the most fundamental questions in this field is: how does CRISPR target the gene of interest? This article delves into the mechanism behind CRISPR’s gene-editing prowess and explains the key steps involved in this groundbreaking process.
The CRISPR-Cas9 system, which is the most widely used CRISPR variant, consists of two main components: the Cas9 protein and a guide RNA (gRNA). The Cas9 protein acts as the “molecular scissors,” while the gRNA serves as the “molecular GPS” that directs the Cas9 protein to the specific location in the genome where the gene of interest is located.
The first step in the process is the design of the gRNA. The gRNA is a short RNA molecule that is complementary to a specific sequence of DNA in the gene of interest. This sequence is known as the target site. The gRNA is designed to be as similar as possible to the target site, with a few mismatches to ensure specificity.
Once the gRNA is designed, it is synthesized and combined with the Cas9 protein. The gRNA binds to the Cas9 protein, forming a complex that can be delivered into the cell. The gRNA guides the Cas9 protein to the target site in the genome.
Upon reaching the target site, the Cas9 protein cuts both strands of the DNA at specific locations flanking the target sequence. This creates a “double-stranded break” in the DNA. The cell’s natural DNA repair mechanisms then come into play.
There are two main pathways for DNA repair: non-homologous end joining (NHEJ) and homology-directed repair (HDR). In the case of CRISPR, the goal is to promote HDR, which is a more precise repair mechanism. To achieve this, a donor DNA template with the desired genetic changes is introduced into the cell along with the CRISPR-Cas9 system.
The cell’s repair machinery uses the donor DNA template as a guide to repair the double-stranded break. This results in the insertion, deletion, or alteration of the target gene sequence, depending on the design of the donor DNA template.
In summary, CRISPR targets the gene of interest by using a gRNA to guide the Cas9 protein to the specific location in the genome. The Cas9 protein then cuts the DNA, and the cell’s repair mechanisms use a donor DNA template to create the desired genetic changes. This innovative technology has the potential to revolutionize medicine, agriculture, and other fields by allowing scientists to edit genes with unprecedented precision and efficiency.