Product Class: Restriction Endonuclease

AlwI

icon_cloned icon_recomb icon_CutSmart dil_A-grey icon_incTemp37 icon_HI_no dam
Alw-I-cutsite_1

Product Introduction

Having supplied restriction enzymes to the research community for over 40 years, NEB has earned the reputation of being the leader in enzyme technologies.  Working continuously to be worth of that distinction, NEB strives to develop enzyme of the highest purity and unparalleled quality. 

  • 100% activity in CutSmart® Buffer (over 210 enzymes are available in the same buffer) allowing for easier double digests
  • Type IIS restriction enzymes recognize asymmetric DNA sequences and cleave outside of their recognition sequence

Catalog # Size Concentration
R0513L 2,500 units 10,000 units/ml
R0513S 500 units 10,000 units/ml

Product Information

Description

Product Source

An E. coli strain that carries the AlwI gene from Acinetobacter lwoffii (R. Morgan).

Reagents Supplied

The following reagents are supplied with this product:

Store at (°C) Concentration
CutSmart® Buffer -20 10 X
Product Categories:
Restriction Endonucleases: A
Applications:
Restriction Enzyme Digestion

Properties and Usage

Unit Definition

One unit is defined as the amount of enzyme required to digest 1 µg of λ DNA (dam-) in 1 hour at 37°C in total reaction volume of 50 µl.

Reaction Conditions

1X CutSmart® Buffer
Incubate at 37°C

1X CutSmart® Buffer
50 mM Potassium Acetate
20 mM Tris-acetate
10 mM Magnesium Acetate
100 μg/ml BSA
(pH 7.9 @ 25°C)

Activity in NEBuffers

CutSmart® Buffer: 100%
NEBuffer 1.1: 50%
NEBuffer 2.1: 50%
NEBuffer 3.1: 10%

Diluent Compatibility

Storage Temperature

-20°C

Storage Conditions

10 mM Tris-HCl
50 mM KCl
1 mM DTT
0.1 mM EDTA
200 μg/ml BSA
50% Glycerol
pH 7.4 @ 25°C

Heat Inactivation

No

Methylation Sensitivity

dam methylation: Blocked
dcm methylation: Not Sensitive
CpG Methylation: Not Sensitive

Notes

  1. For enzymes that cannot be heat-inactivated, we recommend using a column for cleanup (such as the Monarch® PCR & DNA Cleanup Kit), or running the reaction on an agarose gel and then extracting the DNA (we recommend Monarch Gel Extraction Kit), or performing a phenol/chloroform extraction.
  2. This enzyme is blocked by dam methylation. More information can be found at Dam-Dcm and CpG Methylation.
  3. AlwI produces DNA fragments that have a single-base 5´ extension which are more difficult to ligate than blunt-ended fragments. 

Faqs & Tech Tips

FAQs

  1. Is AlwI blocked by dam methylation?
  2. What is the activity of AlwI at 25°C?
  3. Why is AlwI- cut DNA difficult to ligate?
  4. Why is an extended digest not recommended?
  5. How can I increase ligation efficiency?
  6. My restriction enzyme used to be available at a lower concentration. Why does it now come at a higher concentration of 10,000 u/ml?
  7. My enzyme is no longer Time-Saver™ qualified. What happened?
  8. What effect does BSA have on the performance of NEB’s restriction enzymes when included in the new buffers?
  9. How can I access the old NEBuffer Activity Chart?
  10. I tested your restriction enzyme on the substrate DNA recommended by NEB, and it appears to be active, however it does not digest my DNA. What could be the reason?
  11. My restriction enzyme used to work well in the old NEBuffer but the new Performance chart indicates it has lower activity even though the only difference is the addition of BSA and removal of DTT to the new buffers. Why?
  12. Why is my Restriction Enzyme not cutting DNA?
  13. Why do I see a DNA smear on an agarose gel after a restriction digest?
  14. Why do I see additional DNA bands on my gel after a restriction digest?
  15. How many nucleotides do I have to add adjacent to the RE recognition site in order to get efficient cutting?

Tech Tips

This enzyme produces a single base extension, which is difficult to ligate.

Protocols & Manuals

Protocols

  1. Optimizing Restriction Endonuclease Reactions
  2. Double Digest Protocol with Standard Restriction Enzymes

Other Tools & Resources

Selection Charts

Usage Guildelines & Tips

Troubleshooting Guides

Restriction Enzyme Troubleshooting Guide

InteractiveTools

Quality & Safety

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 control's 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.