CRISPR-Cas9 A Promising
Approach For All Genetic
Problems
Presented by:
Shanza Fiaz
BSBOT-05F20
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What is CRISPR Cas9?
History
Components of CRISPR-Cas9
Mechanism
Different Cas proteins and their Functions
Contents Applications of CRISPR-Cas System as
Complex Genetic Approach
Advances in CRISPR-Cas System
Limitations
Future prospects
Conclusion
References
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What is CRISPR-Cas9
Clustered regularly interspaced palindromic repeats (CRISPR)/Cas9 is a gene-
editing technology.
These are the part of Bacterial Immune System which detects and recognizes
foreign DNA.
[Link]
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History
Researchers found CRISPR Sequence in Escherichia coli but did
1987 not characterize their function.
CRISPR Sequence are found to be common in other microbes,.
2000
2002 Coined CRISPR name, defined signature Cas genes.
First experimental evidence foe CRISPR Adaptive Immunity.
2007
First experimental evidence of Cas9 genome in Eukaryotic
2015 and plant cell.
(Zhang et al., 2020) 4
Components of
CRISPR-Cas9
I. Guide RNA
II. Cas9 (CRISPR
Associated Protein 9)
III. Protospacer adjacent
motif (PAM)
IV. CRISPR-RNA (crRNA)
V. Trans-activating
crRNA (tracrRNA)
[Link]
(Redman et al., 2022) 5
Mechanism
CRISPR-Cas system functions in three general steps:
Adaptation or Immunization (involving the acquisition of spacers)
Interference/Biogenesis of CRISPR RNA (CrRNA; encoded by the
repeats-spacer region)
Interference (cleavage of invading nucleic acid)
(Asmamaw & Zawdie 2021)
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Mechanism of CRISPER-Cas9
[Link] 7
Repairing of Double stranded DNA Breaks
[Link] 8
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Applications of CRISPR- •Gene Functions in model
organisms
Cas System As complex •Disease resistance
•Increase in food production
Genetic Approach •Drug production
•Cell-based Therapy
Humans Plants Animals
•Cystic Fibrosis •Nutrient Improvement
•Haemophilia •Adaptation to stress
•Duchenne Muscular Dystrophy •Pathogen resistant
•Lung Cancer •Herbicide resistant
•HIV, Leukemia •Abiotic resistance
•Sickle Cell Anemia •Architecture Asmamaw et al., 2021 10
Tabassum et al., 2023 11
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Recent Advances in CRISPR-Cas9
Technology to enhance plant disease resistant
Knockout of EDR1 gene in wheat which led to the mutant Arabidopsis
plants highly resistant to powdery mildew.
CRISPR-Cs9 edited citrus plants showed resistance to Xanthomonas citri,
the causal agent of citrus bacterial canker.
CRISPR-Cas9 system induce resistance to geminiviruses, beet severe
curly top virus and bean yellow dwarf virus in Arabidopsis and Nicotiana
benthamiana.
(Bortesi & Fischer 2019) 13
Advances in CRISPR-Cas9 for the
Treatment of Muscular Dystrophy
(Tabassum et al., 2023) 14
Use of CRISPR-Cas
Technology in Pakistan
In January 2022, Scientists use CRISPR/Cas9 to improve cotton varieties
against biotic and abiotic stresses. (Shahzad et al.,2022)
CRISPR-Cas system is being engineered to cure COVID-19 patients. Pfizer
and Moderna COVID vaccine build through CRISPR. (Ahmed et al.,2020)
CRISPR-Cas technology is used as therapeutic tool in the reduction and
identification of new tumor target genes in the solid tumors. (Hazafa et
al.,2020)
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Limitations
Limitations
Difficulty of Gene editing at the right cell
Possibility of off-target Effects
Ethical Boundaries
Difficulty of Gene editing at the right
SOLUTION:
cell
Proper selection of
Possibility of off-target
guide RNA Effects
Use smaller guide RNA of 17nt instead of
Ethical Boundaries
20nt.
Sequence the organism first you want to
work with.
Proper Concentration of Cas9 and Length (Asmamaw & Zawdie 2021).
of time for expression of Cas9 16
Agriculture
Drug
Ecology
Discovery
Future
Prospects
Gene
Diagnosis
Therapy
Infectious
Diseases
(Redman et al., 2022)
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CRISPR technology has emerged as a
powerful and universal technology
Conclusion for genome engineering.
This technology has proved its potential by
being user friendly and has shown
its practicality in ensuring health as well as
food security of the future.
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References
Redman, M., King, A., Watson, C., & King, D. (2022). What is CRISPR/Cas9?. Archives of
Disease in Childhood-Education and Practice, 101(4), 213-215.
Zhang, S., Guo, F., Yan, W., Dai, Z., Dong, W., Zhou & Jiang, M. (2020). Recent advances of
CRISPR/Cas9-based genetic engineering and transcriptional regulation in industrial
biology. Frontiers in Bioengineering and Biotechnology, 7, 459.
Asmamaw, M., & Zawdie, B. (2021). Mechanism and applications of CRISPR/Cas-9-mediated
genome editing. Biologics: Targets and Therapy, 353-361.
Tabassum, J., Ahmad, S., Hussain, B., Mawia, A. M., Zeb, A., & Ju, L. (2023). Applications and
potential of genome-editing systems in rice improvement: current and future
perspectives. Agronomy, 11(7), 1359.
Bortesi, L., & Fischer, R. (2019). The CRISPR/Cas9 system for plant genome editing and
beyond. Biotechnology advances, 33(1), 41-52.
Ran, F. A., Hsu, P. D., Wright, J., Agarwala, V., Scott, D. A., & Zhang, F. (2020). Genome
engineering using the CRISPR-Cas9 system. Nature protocols, 8(11), 2281-2308.
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