eSpCas9-GFP Protein

半岛bd体育手机客户端 说明

一般描述

Recombinant enhanced specificity Cas9-GFP protein fromStreptococcus pyogenes(~192 KD) is a ready-to-use reagent for genome engineering experiments. When combined with target-specific guide RNAs, eSpCas9-GFP protein will act as a targeted nuclease suitable for transfection of cell cultures and for the accelerated development of genetically-modified animals via one-cell embryo injection.
An N-terminally fused enhanced green fluorescent protein with an excitation peak at 488 nm and emission peak at 509 nm allows for visualization of transfected RNP complex in addition to utility in flow cytometry applications. The protein also contains three varied nuclear localization sequences postioned for optimal activity.

应用

• Functional Genomics
• Target Validation
• Genome Editing
• Fluorescence Microscopy
• Flow Cytometry

特点和优势

• Enhanced specificity compared to wild type Cas9
• Highly active
• Ready-to-inject/transfect

包装

pkg of 50 μg ( ≥260 pmol )
pkg of 250 μg ( ≥1300 pmol)

组分

Each kit consists of:

• one vial of lyophilized eSpCas9-GFP recombinant protein
• one vial containing 1 mL of 1x dilution buffer
• one vial containing 1 mL of nuclease-free water with glycerol

原理

CRISPR/Cas systems are employed by bacteria and archaea as a defense against invading viruses and plasmids. Recently, the type II CRISPR/Cas system from the bacteriumStreptococcus pyogeneshas been engineered to function in eukaryotic systems using two molecular components: a single Cas9 protein and a non-coding guide RNA (gRNA). The Cas9 endonuclease can be programmed with a single gRNA, directing a DNA double-strand break (DSB) at a desired genomic location. Similar to DSBs induced by zinc finger nucleases (ZFNs), the cell then activates endogenous DNA repair processes, either non-homologous end joining (NHEJ) or homology-directed repair (HDR), to heal the targeted DSB.
Newly engineered eSpCas9 (1.1) enables the efficient targeted gene editing of established CRISPR systems with the benefit of reduced off-target effects. Point mutations (K848A/K1003A/R1060A) in the chromosome-binding motif of SpCas9, as described by Slaymaker, et al., provide higher on-target fidelity without loss of cleavage efficiency.

重悬

Lyophilized enhanced specificityS. pyogeneseSpCas9-GFP protein should be resuspended in the Reconstitution solution provided to desired concentration. Gently tap tube to completely dissolve lyophilized powder, incubate for 10 minutes on ice, and spin tube to bring material to bottom of tube.

其他说明

Use our CRISPR Selection Tool to order gRNA

Check out our other MISSION^?Cas9 Proteins at SigmaAldrich.com/CRISPRproteins

法律信息

CRISPR Use License Agreement
MISSION is a registered trademark of sigma-aldrich Co. LLC

半岛bd体育手机客户端 性质

重组	expressed inE. coli
测定	≥90% (SDS-PAGE)
形式	lyophilized powder
包装	pkg of 1 kit (3 components)
报告基因	GFP
运输	wet ice
储存温度	?20℃

安全信息

储存分类代码	11 - Combustible Solids
WGK	WGK 3