CRISPR-Cas9 Antibody - BSA Free

Novus Biologicals | Catalog # NBP3-05548

Novus Biologicals
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Key Product Details

Species Reactivity

Bacteria

Applications

Western Blot, Immunocytochemistry/ Immunofluorescence

Label

Unconjugated

Antibody Source

Polyclonal Rabbit IgG

Format

BSA Free
View all CRISPR-Cas9 Primary Antibodies »

Product Specifications

Immunogen

N-terminal region, amino acids 1-608 and C-terminal region, amino acids 814-1372 of S. pyogenes CRISPR-CAS9 sequence CDJ55032.1 from S. pyogenes, expressed in and purified from E. coli

Clonality

Polyclonal

Host

Rabbit

Isotype

IgG

Theoretical MW

160 kDa.
Disclaimer note: The observed molecular weight of the protein may vary from the listed predicted molecular weight due to post translational modifications, post translation cleavages, relative charges, and other experimental factors.

Scientific Data Images for CRISPR-Cas9 Antibody - BSA Free

Western Blot: CRISPR-Cas9 Antibody [NBP3-05548]

Western Blot: CRISPR-Cas9 Antibody [NBP3-05548]

Western Blot: CRISPR-Cas9 Antibody [NBP3-05548] - Western blot analysis of HEK293 cell lysates using rabbit pAb to S. pyogenes CRISPR-Cas9 Antibody: [1] protein standard (red), [2] non-transfected cells, [3] transfected cells with C-terminal (814-1372 amino acids) of S. pyogenes CRISPR-Cas9, and [4] transfected HEK293 cells with N-terminal (1-608 amino acids) of S. pyogenes CRISPR-Cas9. 60kDa and 68kDa bands correspond to S. pyogenes CRISPR-Cas9 C-terminal and N-terminal constructs respectively.
Immunocytochemistry/ Immunofluorescence: CRISPR-Cas9 Antibody [NBP3-05548]

Immunocytochemistry/ Immunofluorescence: CRISPR-Cas9 Antibody [NBP3-05548]

Immunocytochemistry/Immunofluorescence: CRISPR-Cas9 Antibody [NBP3-05548] - HEK293 cells were transfected with a construct including the N-terminal 608 amino acids of S. pyogenes CRISPR-Cas9 fused to GFP and stained with CRISPR-Cas9 Antibody in red. Transfected cells express the green fusion protein and bind the antibody in red, producing a yellow signal. Nuclear DNA in transfected and non-transfected cells is revealed with the blue DNA stain DAPI.

Applications for CRISPR-Cas9 Antibody - BSA Free

Application
Recommended Usage

Immunocytochemistry/ Immunofluorescence

1:1000-1:2000

Western Blot

1:1000-1:10000

Formulation, Preparation, and Storage

Purification

Immunogen affinity purified

Formulation

50% PBS, 50% glycerol

Format

BSA Free

Preservative

5mM Sodium Azide

Concentration

1 mg/ml

Shipping

The product is shipped with polar packs. Upon receipt, store it immediately at the temperature recommended below.

Stability & Storage

Store at 4C short term. Store at -20C long term. Avoid freeze-thaw cycles.

Background: CRISPR-Cas9

Clustered regularly interspaced short palindromic repeats (CRISPRs) are derived from DNA fragments of bacteriophages that infect prokaryotes. When infected, the bacteria capture snips of DNA from the invading virus to create CRISPR arrays. During subsequent infections, the bacteria produce RNA segments from the CRISPR arrays to target the virus' DNA. CRISPR-associated protein 9 (Cas9) is RNA-guided, binds DNA, and is a cleaving enzyme that functions as an integral component of the bacterial CRISPR adaptive immune system that targets the virus' DNA to disable it (1). To check for sites complementary to the 20 base pair spacer region of the guide RNA (gRNA) of the CRISPR, Cas9 unwinds foreign DNA that invades the bacteria. If the DNA substrate is complementary to the gRNA, Cas9 cleaves the invading DNA, rendering the virus disabled. The presence of a 5'-NGG-3' protospacer adjacent motif (PAM) sequence immediately downstream of the target DNA (protospacer) is required for Cas9 cleavage of foreign DNA. As PAM is absent in bacterial CRISPR loci, cleavage of the host genome is avoided and provides a novel sequence for identification of foreign DNA by Cas9.

Using CRISPR-Cas9 technology, double-stranded DNA breaks may be induced within specific targeted genome sequences (target DNA; protospacer) for insertion or removal of DNA sequences for gene editing applications. To target a specific loci, a gRNA that will bind to a specific target sequence of DNA within a genome is created. The gRNA will recognize the DNA sequence, and the Cas9 enzyme will cleave the DNA at the targeted location. Once the targeted DNA is removed by Cas9, the cell's own DNA repair mechanism is used to insert or remove a DNA sequence for genomic editing.

Cas9 detection is used to confirm and evaluate CRISPR Cas9 gRNA transfection efficiency. Western blot analysis of CRISPR-Cas9 gRNA transfected cell lysates with Cas9 antibodies identifies the protein having a theoretical molecular weight of 160kDa. Broad areas of research are benefiting from CRISPR-Cas9 based gene editing tools including studies of basic immunity functions, genetic screening and disease treatment (2). Ethical concerns have led to many countries making it illegal to manipulate human germline cells or perform embryo genome editing.

References

1. Oakes, B. L., Fellmann, C., Rishi, H., Taylor, K. L., Ren, S. M., Nadler, D. C.,... Savage, D. F. (2019). CRISPR-Cas9 Circular Permutants as Programmable Scaffolds for Genome Modification. Cell, 176(1-2), 254-267.e216. doi:10.1016/j.cell.2018.11.052

2. Chiou, S. H., Winters, I. P., Wang, J., Naranjo, S., Dudgeon, C., Tamburini, F. B.,... Winslow, M. M. (2015). Pancreatic cancer modeling using retrograde viral vector delivery and in vivo CRISPR/Cas9-mediated somatic genome editing. Genes Dev, 29(14), 1576-1585. doi:10.1101/gad.264861.115

Long Name

CRISPR-associated Protein 9

Alternate Names

Cas9, CRISPR-associated endonuclease Cas9/Csn1, CRISPR-Cas9/Csn1, CRISPR/Cas9, csn1, SPy_1046, SPy1046, SpyCas9

Additional CRISPR-Cas9 Products

Product Documents for CRISPR-Cas9 Antibody - BSA Free

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Product Specific Notices for CRISPR-Cas9 Antibody - BSA Free

This product is for research use only and is not approved for use in humans or in clinical diagnosis. Primary Antibodies are guaranteed for 1 year from date of receipt.

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Protocols

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