Product Class: Restriction Endonuclease

PaqCI®
neb31 cloned at NEB recombinant 2site dil_B 37 65 Heat CpG

PaqCI has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10211228. Learn more.

Learn about Ligase Fidelity and Push the Limits of Golden Gate Assembly (50+ fragments).

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.


Catalog #R0745

NEB restriction endonuclease that recognizes the sequence TTA_AT^TAA
Isoschizomers | Single Letter Code

Product Introduction

Product Information

Description

PaqCI digests to completion and does not exhibit star activity



1 µg Lambda DNA was digested with 8 units of either PaqCI (NEB #R0745) or AarI (Thermo Fisher) following manufacturer’s recommended protocols. Digestions were analyzed on a 1% agarose gel. The results indicate that unlike AarI, PaqCI cuts to completion and does not exhibit star activity.

Product Source

An E. coli strain that carries the modified cloned PaqCI gene from Paucibacter aquatile.    
This product is related to the following categories:
Restriction Endonucleases P R Products
This product can be used in the following applications:
High-throughput cloning and automation solutions,
NEBridge® Golden Gate Assembly

Properties & Usage

Unit Definition

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

Reaction Conditions

1X rCutSmart™ Buffer
Supplement with PaqCI Activator
Incubate at 37°C

rCutSmart™ Buffer
50 mM Potassium Acetate
20 mM Tris-acetate
10 mM Magnesium Acetate
100 µg/ml Recombinant Albumin
(pH 7.9 @ 25°C)

Activity in NEBuffers

NEBuffer™ r1.1: 10%
NEBuffer™ r2.1: 100%
NEBuffer™ r3.1: 10%
rCutSmart™ Buffer: 100%

Diluent Compatibility

Storage Buffer

10 mM Tris-HCl
300 mM NaCl
1 mM DTT
500 µg/ml Recombinant Albumin
50% Glycerol
pH 7.4 @ 25°C

Heat Inactivation

65°C for 20 minutes

Methylation Sensitivity

dam methylation: Not Sensitive
dcm methylation: Not Sensitive
CpG Methylation: Impaired by Overlapping

Product Notes

  1. PaqCI is an isoschizomer of AarI.
  2. This enzyme is impaired by CpG methylation. More information an be found at "Dam-Dcm and CpG Methylation."
  3. PaqCI is an enzyme that requires multiple recognition sites to achieve complete digestion.  NEB has therefore included “PaqCI Activator” which can be added to the reaction with 1:1 ratio (i.e. 1 ul enzyme to 1 ul PaqCI Activator).  Please note that for Golden Gate Assembly reactions, the amount of PaqCI Activator varies depending on how many fragments are being assembled. 
  4. star activity may result from extended digestion, high enzyme concentration or a glycerol concentration of >5%

Protocols, Manuals & Usage

Protocols

  1. Golden Gate Assembly Protocol using PaqCI® (NEB #R0745) and NEBridge Ligase Master Mix (NEB #M1100)
  2. Golden Gate Assembly Protocol using PaqCI (NEB #R0745) and T4 DNA Ligase (NEB #M0202)

Usage & Guidelines

Tools & Resources

Selection Charts

Web Tools

FAQs & Troubleshooting

FAQs

  1. Which restriction enzymes are used in Golden Gate Assembly?
  2. Which restriction enzymes are used in GoldenBraid Assembly?
  3. When is star activity a concern?
  4. Why is PaqCI not cutting DNA to completion?
  5. Can you tell me more about the switch from BSA to Recombinant Albumin (rAlbumin) in NEBuffers?

Troubleshooting

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.

Specification Change Notifications

PaqCI has been reformulated with Recombinant Albumin (rAlbumin) beginning with Lot #10211228. All subsequent (higher number) lots will contain rAlbumin. For additional information, please visit at NEB Restriction Enzyme formulations with Recombinant Albumin (rAlbumin).

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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