An artistic illustration of the use of CRISPR in genetic editing
What is CRISPR?
CRISPR, an abbreviation for "clustered regularly interspaced short palindromic repeats," is a tool that enables the precise alteration of DNA in living organisms.
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Originating from naturally existing genome editing mechanisms within bacteria, CRISPR was adapted from the bacterial immune system and integrated into laboratory use.
How does CRISPR work?
1. Bacterial defense system
Bacteria have a natural defence mechanism against viruses, utilising a process known as the CRISPR array, where DNA segments from viruses are integrated into the bacterial DNA when the virus enters the bacteria, thus preserving a "mugshot" of such viruses.
3. Utilisation
Inspired by this mechanism, scientists have recently repurposed the CRISPR-Cas system to enable precise gene editing in a wide range of organisms, opening doors to various applications in medicine, agriculture, and environmental conservation (elaborated more in the application section).
2. The Cas Enzyme
Because of this record, upon being attacked by a previously encountered virus, the bacteria is able to recognise this virus and employ an enzyme referred to as a CRISPR-associated protein (Cas) to locate and cut the viral DNA, effectively stopping its further attacks.
4. Cas9
One of the Cas enzymes, Cas9, exhibits the capability to accurately cleave DNA strands. Due to its capacity for swift, straightforward, and precise gene editing across numerous plants and animals, Cas9 is frequently employed in CRISPR processes involving a diverse array of organisms.
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The detailed process of CRISPR
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Please click on the image for larger, clearer diagrams as well as explanations and labels provided along the diagrams.
A copy of the target DNA sequence that scientists were trying to remove/alter was provided. This is called the "guide RNA" and will later bind to the Cas9 enzyme.
To direct the Cas9 enzyme (dark green on the diagram) to their target sequence, we attach them to the synthetic guide (purple on the diagram). Since the guide was a copy of the DNA segments that scientists are trying to alter/remove, by using this as a reference, the Cas9 enzyme would know where to cut.
The guide and Cas9 look for a match to the guide.
Once the sequence that matches that of the guide's was found, Cas9 cut through it.
The DNA segments that the scientists were trying to remove are now removed.
The cell could repair the cut by joining the broken ends together.
The cell could also repair the cut using available DNA nearby.
Applications of CRISPR
CRISPR technology boasts a diverse range of applications across fields such as medicine, agriculture and environmental conservation.
Its precise gene-editing capabilities have revolutionized research and development in these areas, offering solutions to various challenges and driving innovation in scientific endeavors.
Therapeutic Applications
The most prominent application of CRISPR lies in its ability to repair mutations that cause diseases, thereby influencing gene expression and opening up new treatment options. CRISPR therapy trials are ongoing for ailments such as cancer, blood disorders, and hereditary blindness. Initial studies using CRISPR-Cas9 for gene editing in individuals with sickle cell disease and beta-thalassemia have yielded encouraging outcomes, indicating the effectiveness of CRISPR methods in therapy.
Agriculture
CRISPR-based genome editing offers exciting opportunities for improving crop characteristics, including yield, pest and disease resistance, while also reducing agriculture's environmental impact. This is exemplified by a recent study that showcased the application of CRISPR-Cas9 in developing disease-resistant rice plants, which increased in yield without needing additional input of human effort, underscoring the potential of CRISPR-based strategies for crop enhancement. These innovations hold promise for addressing worldwide concerns regarding food security and sustainability.
Biofuel production
CRISPR technology also holds potential for enhancing the efficiency and sustainability of biofuel production by refining the traits of biofuel feedstock organisms. For example, scientists have utilised CRISPR to increase the lipid content and improve tolerance to environmental stress in microalgae species commonly used for biofuel production.
Advantages & Benefits
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Precision + Accuracy
CRISPR technology provides a meticulous approach to pinpointing genetic sequences, empowering scientists to manipulate DNA at exact sites within the genome with exceptional precision. This accuracy diminishes the risk of off-target, thereby amplifying the safety and effectiveness of genetic manipulation techniques.
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Versatility + Flexibility
CRISPR systems have a wide range of applications, including gene correction, activation, and repression. Furthermore, CRISPR-derived instruments have the capacity to address both DNA and RNA as well as modify multiple genes within one cell simultaneously. This broadens their applicability in multiplexed gene editing, genetic control, and gene manipulation.
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Efficiency + Scalability
CRISPR-mediated gene editing proves highly efficient, facilitating quick and uncostly modification of genetic material. Furthermore, CRISPR technologies are easily scalable for large-scale applications. Particularly noteworthy is CRISPR-Cas9, which stands out as a budget-friendly gene editing method, delivering results within significantly shorter timeframes (several days) compared to conventional approaches that may span months or years. This accelerated pace increased scientific progress and enhances research efficiency.
Disadvantages
Despite all of the aforementioned advantages, CRISPR has its drawbacks and limitations.
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Off-target effects happen when Cas enzymes unintentionally modify DNA at locations different to the intended target, which can result in the risk of undesirable genetic mutations. This phenomenon can't be completely avoided even for highly specific CRISPR-Cas systems, underscoring the critical drawback of CRISPR..
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Utilizing CRISPR technology evokes ethical considerations concerning the possible misuse of genome editing, encompassing issues such as the creation of "designer babies", changes to the germline with uncertain long-term implications, and the exacerbation of preexisting social disparities.
An image demonstrating the possibility of Cas enzymes effecting off-target sections of the DNA sequence
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Mosaicism describes the existence of genetically distinct cell populations within an organism, often stemming from incomplete or imperfect editing by CRISPR-Cas systems. Recent studies have shown that CRISPR-mediated genome editing in early-stage embryos can lead to mosaicism, where distinct cells exhibit various genetic modifications. This phenomenon raises concerns regarding the consistency and predictability of editing outcomes.
Cost of CRISPR
Providing a definitive assessment of whether CRISPR technology is economically viable is challenging. While certain aspects of genome editing technology demonstrate cost-effectiveness, inherent limitations and constraints can increase associated expenses.
Reasons for CRISPR to be considered cheap
Reasons for CRISPR to be considered expensive
ease of use
CRISPR offers simplified genome editing compared to older methods like TALENs or Zinc Finger Nucleases (ZFNs). Its accessibility benefits researchers of varying expertise and resource levels, allowing them to work effectively.
cheap reagents
CRISPR experiments benefit from the affordability of essential components like Cas proteins and guide RNAs, which are commercially available at relatively low costs. This enables researchers to conduct experiments economically.
equipments
Performing CRISPR experiments relies on specialized laboratory equipment and infrastructure, such as PCR machines and cell culture facilities. Acquiring and maintaining this equipment substantially adds to the overall costs involved.
R&D cost
CRISPR technology required substantial upfront investment in research and development (R&D). Scientists spent years characterizing CRISPR-Cas systems and improving editing tools. These initial costs are usually reflected in the pricing of commercial CRISPR products and services.
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