This document summarizes a presentation on CRISPR/Cas genome editing. It defines CRISPR/Cas as a type of genetic engineering that uses artificially engineered nucleases to make specific cuts in DNA. It describes the CRISPR/Cas system's origins and components, including Cas9, guide RNA, and PAM sequences. Applications discussed include genome editing in animals and plants, as well as concerns over off-target effects. Companies offering CRISPR services or kits are also mentioned.

1. S. Mohammadpour
PhD student in Molecular Genetic
Supervised by: Dr. Goodarzi
University of science and research
Shiraz Iran
January 2016

2. Outlines:
1. Definition
2. CRISPR/Cas system
3. Applications
4. News and concerns
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Genome editing, or genome editing with engineered
nucleases (GEEN)
•Is a type of genetic engineering
•DNA is inserted, replaced, or removed from a genome
•Uses artificially engineered nucleases, or "molecular scissors."
•Creates specific double-strand breaks (DSBs) at desired locations
in the genome
•Cell’s endogenous mechanisms repairs the induced break by
(HDR) and (NHEJ)
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4. 365 days: Nature’s 10
Ten people who mattered this year.
18 December 2013
FENG ZHANG: DNA’s master editor
Borrowing from bacteria, a biologist helps to create a
powerful tool for customizing DNA.
By Daniel Cressey
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About Feng Zhang…
Born: 1982
Nationality: American
Fields: Neuroscience, Bioengineering
Institutions: MIT, Broad Institute, Alma mater, Harvard University,
Stanford University
Academic advisors: Karl Deisseroth
Known for: Optogenetics, CRISPR
Notable awards: Alan T. Waterman Award, National Institutes of
Health Director's Pioneer Award, Perl-UNC Neuroscience Prize,
Jacob Heskel Gabbay Award in Biotechnology and Medicine,
Tsuneko & Reiji Okazaki Award
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7. • First described (1987)
• 48% bacteria; 84% archaea
•Short Regularly Spaced Repeats (SRSR) (2000)
• Renamed CRISPR (2002)
• A prokaryotic adaptive immune system (2005-12)
• Identify Cas 9 (2012)
• Genome Engineering by CRISPR/Cas system (2013)
CRISPR
Clustered Regularly Interspaced Short Palindromic Repeats
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8. CRISPR/Cas9 nomenclature
CRISPR: Clustered Regularly Interspaced Palindromic Repeats
Cas9: CRISPR associated protein 9
a nuclease, an enzyme specialized for cutting DNA Cas1..Cas10 exist
gRNA: guide RNA
a construct/chimera of CRISPR RNA (crRNA) and trans-activating CRISPR RNA
(tracrRNA)
PAM: protospacer adjacent motif with sequence
NGG (any, guanine, guanine)
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9. CRISPR/Cas mechanism in bacteria
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10. CRISPR/CAS Programmable DNA scissor
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11. Delivery to cells in humans
lentivirus vectors
Via stem cells
Via liposomes
Off-target effects and
unintended DNA cleavage
paired nickases
(single-strand DNA break)
double-nicking
Check with whole genome sequencing
modified Cas9 variants
paired nicking
modified sgRNAs
Variable DNA cleavage
efficiencies
CRISPER/CAS Problems
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12. Application examples
Source: Hsu, Lander, Zhang: Development and Applications of CRISPR-Cas9 for Genome
Engineering; Cell 157, June 5, 2014
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Application examples
Genome-wide inactivation of porcine endogenous retroviruses (PERVs)
George Church, Department of Genetics, Harvard Medical School, Boston, MA, USA.Oct
2015.
Generation of gene -target dogs using CRISPR/Cas9 system
Xiang Gao, CAS Key Laboratory of Regenerative Biology, South China Institute for Stem
Cell. Oct 2015.
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14. Organisms that have been modified using
the CRISPR–Cas9 system
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CRISPR in Zhang Lab
1-Development and Applications of CRISPR-Cas9 for Genome Engineering. (2014).
2-Improved vectors and genome-wide libraries for CRISPR screening. (2014).
3-Efficient CRISPR-Cas9-mediated genome editing in Plasmodium falciparum.Wagner. (2014).
4-CRISPR-mediated direct mutation of cancer genes in the mouse liver. (2014).
5-CRISPR-Cas9 Knockin Mice for Genome Editing and Cancer Modeling. Platt. (2014)
6-In vivo interrogation of gene function in the mammalian brain using CRISPR-Cas9. (2014).
7- Genome Engineering Using CRISPR-Cas9 System. (2014).
8- Genome-scale transcriptional activation by an engineered CRISPR-Cas9 complex. (2014).
9- Genome-wide CRISPR Screen in a Mouse Model of Tumor Growth and Metastasis. (2014).
10-CRISPR germline engineering-the community speaks. (2015).
11- High-throughput functional genomics using CRISPR-Cas9. (2015).
12- CRISPR/Cas9 cleavage of viral DNA efficiently suppresses hepatitis B virus. (2015).
13- Sequence determinants of improved CRISPR sgRNA design. (2015).
14- CRISPR-Cas9: Prospects and Challenges. (2015).
15- A Genome-wide CRISPR Screen in Primary Immune Cells to Dissect Regulatory Networks.
(2015).
16- BCL11A enhancer dissection by Cas9-mediated in situ saturating mutagenesis. (2015).
17- Cpf1 Is a Single RNA-Guided Endonuclease of a Class 2 CRISPR-Cas System. (2015).
18- Orthogonal gene knockout and activation with a catalytically active Cas9 nuclease. (2015).
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Baby Design Companies
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17. The future of CRISPR/Cas9
Doudna J and Charpentier E; ScienceMag 2014 .
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18. Software and databases
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19. Companies offering CRISPR/Cas9 services
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20. Companies
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Do-it-yourself CRISPR genome editing kits bring
genetic engineering to your kitchen bench
• LOZ BLAIN, NOVEMBER 11, 2015
The Open Discovery Institute
(ODIN)
Josiah Zayner
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“The Human SPIRIT Has No Gene”
THANK YOU!S.Mohammadpour