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LIVE – Now –>> CRISPR-Cas9 Discovery and Development of Programmable Genome Engineering – Gabbay Award Lectures in Biotechnology and Medicine – Hosted by Rosenstiel Basic Medical Sciences Research Center, 10/27/14 3:30PM Brandeis University, Gerstenzang 121

Reporter: Aviva Lev-Ari, PhD, RN

REAL TIME Conference Coverage by Dr. Aviva Lev-Ari, PhD, RN

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LIVE CONTENT from 

Gabbay Award Lecture in Biotechnology and Medicine @Brandeis

is been added 0n 10/27 @3;30PM

InVivo Target Search Mechanism – Fast Cas9 Diffusion in Live Cells

First Speaker

Emmanuelle Charpentier (Dept. of Molecular Biology. Umea University; Dept. of Regulation in Infection Biology, Helmholtz Centre for Infection Research)

CRISPR-Cas9: How a Bacterial Immune System Revolutionizes Life Sciences and Medicine

application in dairy industry basilicum, bacteria

Second Speaker

Jennifer Doudna (Dept. of Biochemistry, Biophysics and Structural Biology, UC Berkeley)CRISPR-Cas9 Discovery and Development of Programmable Genome Engineering

Bacterial/archaeal chromosome

a single gene CAs9m- enzyme single-guide RNAs (sgRNAs)

crRNA

A Programmable Dual-RNA- Genome editing begins with dsDNA cleavange

ZFHs, TALENs, HEs, Cas9:sgRNA

protein DNA recognition

Genome targeting technologies: ZFN & TALEN

/Cas9/targeting RNA bound by a nuclease

2012-RNA-guided DNA endonuclease

1/2013 Church, Zhang

Re-Writing the Genome:

DNA structu

restriction enzymes

PCR

Specific genome editing in cell and in organizmism

robust transcriptional control with ccatalyrtic

In the Lab @Berkeley

Hoe Cas9 find DNA targets with high specificity

Mechanism of DNA interrogation

• high affinity product binding, no substrate turnover
• binding first occur at PAM motifs
• PAM binding triggers Cas9 catalytic activity

CRISPR: Clusters of Regulary

three steps to acquire immunity in bacteria

1. adaptation

2. crRNA biogenesis

3. Interference

CRISPR in Structural Biology

• RNA-induced conversion of Cas9 into an active confrontation – crystal structure morphology

Models for DNA interrogation by Cas9:RNA

Programmed RNA cleavage using Cas9:gRNA in O’Connell at al. (2014) Nature

 

• Therapeutic Applications

1. Delivery of antibiotic small molecule

2. Animal models live cell (catalitic inactive version vs active version)

 

Third Speaker

Feng Zhang (McGovern Inst. for Brain Research, MIT)
Development and Applications of CRISPR-Cas9 for Genome Editing

Brain Research – Applications for CRISPER

• Genetics & epigenetics
• signals – optogenetics Neuromodulation
• optogenetics — fibers — cells

The CRISPR/Cas bacterial

Crispr RNA maturation

E.coli – immunity for using Crispr

Streptococcus thermophilus – CRISPr editing applied

Development of a Technology Platform

• expand modes of Genomic Pertubation

develop efficient co-transduction of primary neurons

multiplex knockout un the denate

• demonstrate uses in biological therapeutic context

CRISPR — Reagents development, protocol developed, discussion forums

targeting of MeCP2 gyrus leads to robust protein depletion and behavior change

toxicity potential researched

Cre-dedendent CAs9 MOUSE FOR CANCER MODELING – mouth lung

• develop an open source for the community

Genome-scale CRISPR knockout (GeCKO) Screen – generate library pool of cell – mutation identification – melanoma treated by BRAF inhibitor – targets for menanoma resistence

Comparison of shRNA vs GeCKO – CRISPR more robust and statisticaly significant, However, GeCKO provides higher sensitivity than shRNA

•  foundational technology development

Crystal structure of Cas9 in complex wiht guided RNA and target DNA

Cre-dependent Cas9 Mouse

 

For Twitter.com

#Cas9
#CRISPR
#molecularbiology
#biotechnology
#GeneEditing
#genetic#engineering (sometimes these two # are put together like this)
#Boston

@BrandeisU
@MIT
@UCBerkeley
@CRISPRpapers
@UmeaUniversity
@mcgovernmit (this is for Feng Zhang)
@pharma_BI

 

CRISPR Service / Cas9

Commercialization by

 http://www.appliedstemcell.com/services/cell-line-models/cell-line-modification/

The CRISPR/Cas9 system uses the Cas9 nuclease to facilitate RNA-guided site-specific DNA cleavage. The system consists of two components:

(1) Mammalian codon-optimized version of the Cas9 protein carrying a nuclear localization signal to ensure nuclear compartmentalization in mammalian cells

(2) Guide RNAs (gRNAs) to direct Cas9 protein to sequence-specifically cleave the targeted DNA

The advantage of CRISPR/Cas9 over ZFNs or TALENs is its scalability and multiplexibility in that multiple sites within the mammalian genome can be simultaneously modified, providing a robust, high-throughput approach for gene editing in mammalian cells.

CRISPR Service (Point mutation, Deletion, Small DNA insertion)

We are experts in CRISPR Service! Applied StemCell is in Nature Biotechnology as one of the select companies for CRISPR-Cas9 tools! Monya Baker, “Gene Editing at CRISPR Speed,” Nature Biotechnology, 32: 309-312, April 2014.

Since Applied Stem Cell started out as a company focused on induced pluripotent stem cells (iPSCs), we have excellent capabilities of correcting mutations in disease-model iPSCs using CRISPR/ Cas9.