Product Class: Other

EnGen® Sau Cas9
neb31 recombinant 37 65 Heat

The S size of this product was discontinued on July 31, 2023. M0654T will remain available for purchase.

We are excited to announce that all reaction buffers are now BSA-free. NEB began switching our BSA-containing reaction buffers in April 2021 to buffers containing Recombinant Albumin (rAlbumin) for restriction enzymes and some DNA modifying enzymes. Find more details at www.neb.com/BSA-free.

Product Introduction

EnGen® Sau Cas9 is a programmable DNA endonuclease guided by a single guide RNA (gRNA).  Targeting requires gRNA complementarity to the target site as well as a 5′NNGRRT protospacer adjacent motif (PAM) on the DNA strand opposite the target sequence.  Cleavage by EnGen® Sau Cas9 occurs ~3 bases upstream of the PAM and leaves blunt ends.

  • Longer 5′ NNGRRT 3′ PAM sequence allows for higher specificity
  • Two nuclear localization signals for improved transport into the nucleus
  • High concentration liquid format option can be used for microinjection, electroporation and lipofection
  • Compatible with the EnGen® Mutation Detection Kit
  • Need guide RNA? NEB recommends the Horizon™ CRISPR Design Tool for designing and ordering synthetic guide RNA for EnGen Sau Cas9
Catalog # Size Concentration
M0654S 90.0 pmol 1 µM
M0654T 500.0 pmol 20 µM

Product Information

Description

EnGen® Sau Cas9 from Staphylococcus aureus is an RNA-guided DNA endonuclease that catalyzes site-specific cleavage of double-stranded DNA.  Targeting requires a ~100 nucleotide guide RNA (gRNA) complementary to the target site, as well as a 5′ NNGRRT 3′ (N = any, R = A or G) protospacer adjacent motif (PAM) on the DNA strand opposite the target sequence.  DNA cleavage by EnGen Sau Cas9 occurs ~3 bases 5′ of the PAM.  EnGen Sau Cas9 has Simian virus 40 (SV40) T antigen nuclear localization signals (NLS) at both the N- and C-termini of the protein.

Schematic representation of Sau Cas9 nuclease sequence recognition and DNA cleavage



Product Source

EnGen® Sau Cas9 from Staphylococcus aureus is expressed as a C-terminal 6X His-tagged fusion in E. coli.
This product is related to the following categories:
Genome Editing Products,
Programmable Nucleases Products,
This product can be used in the following applications:
Genome Editing Applications

Reagents Supplied

Reagents Supplied

The following reagents are supplied with this product:

NEB # Component Name Component # Stored at (°C) Amount Concentration

Properties & Usage

Reaction Conditions

1X NEBuffer™ r3.1
Incubate at 37°C

NEBuffer™ r3.1
100 mM NaCl
50 mM Tris-HCl
10 mM MgCl2
100 µg/ml Recombinant Albumin
(pH 7.9 @ 25°C)

Storage Buffer

20 mM Tris-HCl
300 mM NaCl
0.1 mM TCEP
50% Glycerol
pH 7.5 @ 25°C

Heat Inactivation

65°C for 5 minutes

Product Notes

  1. 20 µM is equal to ~2.54 mg/ml EnGen® Sau Cas9.  1 µM is equal to ~0.127 mg/ml.

Protocols, Manuals & Usage

Protocols

  1. How to synthesize a sequence-specific sgRNA for experiments using EnGen® Sau Cas9

FAQs & Troubleshooting

FAQs

  1. How can the genome editing efficiency be determined following EnGen® Sau Cas9 (NEB #M0654) transfections or microinjections?
  2. Which nuclear localization signal is fused to EnGen® Sau Cas9?
  3. Can the EnGen® sgRNA Synthesis Kit (NEB #E3322S) be used to synthesize sgRNAs specific for Sau Cas9?
  4. What is the sequence for a typical guide RNA specific for EnGen® Sau Cas9?
  5. How can I synthesize a sequence-specific sgRNA for experiments using EnGen® Sau Cas9?
  6. Why do I observe incomplete digestion with EnGen® Sau Cas9 and my target-specific Sau sgRNA in vitro?
  7. How do I dilute the enzyme to 1 μM for in vitro reactions?
  8. Can you tell me more about the switch from BSA to Recombinant Albumin (rAlbumin) in NEBuffers?

Quality, Safety & Legal

Quality Assurance Statement

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

Specifications

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

Certificate Of Analysis

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

Safety DataSheets

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

Legal and Disclaimers

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally developed. The use of this product may require the buyer to obtain additional third-party intellectual property rights for certain applications. For more information, please email busdev@neb.com.

This product is intended for research purposes only. This product is not intended to be used for therapeutic or diagnostic purposes in humans or animals.

New England Biolabs (NEB) is committed to practicing ethical science – we believe it is our job as researchers to ask the important questions that when answered will help preserve our quality of life and the world that we live in. However, this research should always be done in safe and ethical manner. Learn more.

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