Research Case Study
By: Tanvi Mareddy
Introduction
The CRISPR system is a natural defense mechanism in prokaryotic organisms (bacteria and archaea) and operates through CRISPR arrays with spacers (foreign DNA sequences) and palindromic repeats (DNA sequences that read the same forward and backward). The spacer sequences act as genetic memory while the palindromic repeats act as recognition sites for Cas9 protein. In the process, the CRISPR array is transcribed into pre-crRNA. TracrRNA then helps convert the precursor molecule into the mature guide RNA.
The Cas9 nuclease protein cuts DNA at precise locations while guide RNA recognizes and binds to specific DNA sequences. When the microorganisms encounter foreign genetic material, the CRISPR system captures parts of the invader's DNA and integrates them into its genome as spacer sequences. If the immune system is attacked by the same invader, the guide RNA identifies the specific DNA sequence by matching it with the stored spacer sequences. Subsequently, the Cas9 protein cuts the viral DNA, preventing replication and causing disruption.
History of CRISPR Technology
A historical breakthrough in genetic engineering occurred when scientists figured out CRISPR RNA (crRNA) was a different entity than trans-activating CRISPR RNA (tracrRNA). Scientists were able to synthesize these two entities into a single guide RNA (sg-RNA) in labs. When the sg-RNA and a cas9 protein are paired, they form an effector complex, which allows scientists to target and cleave any sequence of 20 nucleotides.
Homology-directed repair (HDR) and Non-Homologous End Joining (NHEJ) are used as the two repair mechanisms. HDR uses a provided homologous template to repair the double-strand break. This allows for accurate modifications, insertions, or replacements. NHEJ directly joins the broken ends, introducing small insertions or deletions (indels). HDR is employed for precise genome modifications, while NHEJ is error-prone and used for inducing random mutations or gene disruptions.
Applications
CRISPR as a method of genome editing is still relatively new, however, future implications continue to be explored. CRISPR editing in Cancer research and treatment could help us edit DNA within cancer cells and find underlying cancer development quicker.CAR-T cell therapy is when you modify a patient’s T cells to target and attack cancer cells. CRISPR technology can help us edit these T cells in a more efficient and accurate manner.
Citations:
“Car T Cells: Engineering Immune Cells to Treat Cancer.” National Cancer Institute, www.cancer.gov/about-cancer/treatment/research/car-t-cells. Accessed 15 Jan. 2024.
YouTube, YouTube, 29 Oct. 2021, https://www.youtube.com/watch?v=IiPL5HgPehs. Accessed 15 Jan. 2024.
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