3. 3
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)
S.Mohammadpour
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
4
S.Mohammadpour
5. 5
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
S.Mohammadpour
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
11
S.Mohammadpour
12. Application examples
Source: Hsu, Lander, Zhang: Development and Applications of CRISPR-Cas9 for Genome
Engineering; Cell 157, June 5, 2014
12
S.Mohammadpour
13. 13
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.
S.Mohammadpour
14. Organisms that have been modified using
the CRISPR–Cas9 system
14
S.Mohammadpour
15. 15
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).
S.Mohammadpour
23. 23
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
S.Mohammadpour