CRISPR Technology
Genome editing technology has always been a challenging area, which holds great promise for genome manipulation and gene therapy. Conventionally, zinc-finger nucleases and transcription activator-like effector nucleases were used for genome editing. Over the last few years, CRISPR (clustered regularly interspaced short palindromic repeat) restoration system is considered as one of the most significant players in medical genetic research, and this technology has appeared as a versatile and convenient (epi)genome editing tools. CRISPR and CRISPR-associated (Cas) proteins such as catalytically inactivated Cas9 (dead Cas9, dCas9) and scaffold-incorporating single guide sgRNA (scRNA) are found in the genomic loci in bacteria and archaea where it functions as an adaptive the immune system thus targeting and destroying genetic parasites at the DNA level. CRISPR-Cas9 and its variants hold the potential of transforming various genomic screen studies in the upcoming future. Bacteria can be protected from repeated viral attacks by the CRISPR immune system via three steps- adaptation, production of CRISPR RNA, and targeting. In the adaptation phase, DNA from an attacking virus is merged into the CRISPR locus as new spacers of the bacterial genome. In the second step, CRISPR repeats and spacers in the bacterial DNA is transcribed into a short piece of RNA called a CRISPR RNA (crRNA). crRNA is exact matches to the viral genome and thus serve as excellent guides. Finally, crRNA guides the Cas-RNA complex to destroy the viral material by invading bacteriophage DNA via Watson-Crick base pairing. CRISPR technology has potential applications including-
- Used in knockout, knockdown, and activation screens
- Used for targeting coding and non-coding regions throughout the genome
- Eliminating a gene without interfering with intracellular mechanisms
- Identify the performance of defective genes in many chronic diseases
CRISPR/Cas9 technology can be used for developing a sustainable and prolific agricultural system by producing site-specific mutagenesis or targeting transcriptional regulation. Genome editing at target sites by specific nucleases can improve yield, abiotic stress tolerance, enhance resistance to diseases and pests and modify plants for product quality. Moreover, this technology can add new and unique characteristics within a short period of time and is used for the improvement of crop plants in tropical climates. Still, the function of a various number of genes remains unknown, therefore, further efforts are needed to optimize the CRISPR/Cas9 protocols in order to have a greater impact on agriculture in tropical areas. With a rising incidence globally, cancer is one of the main causes of disease-associated mortality and remains a major economic and social burden. CRISPR/Cas9 has become a powerful method to expedite cancer research by dissecting chemical-genetic interactions, manipulating non-coding regions of the genome, providing insight into how tumors respond to drug treatment, discovering novel targets for cancer therapy, and elimination or inactivation of carcinogenic viral infections. This system can be advantageous for industrial processes that utilize bacterial cultures to improve culture sustainability and lifespan. The most important CRISPR applications are-
- Pathogen diagnostics
- Cancer screening
- Metabolic engineering and manufacturing of high-value compounds
- Mouse models or gene deletions
- Correction of deleterious mutations in embryos
- CRISPR genome editing for improved animal breeding
- Gene drives to eradicate malaria
- Modification of traits in living organisms
- Cell therapy
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