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Rapid PCR Cleanup Enzyme Set

recombinant incubation temp heat inactivation
Catalog # Size
E2622 S 100 reactions
E2622 L 500 reactions
E2622 X 2000 reactions
E2622 E 5000 reactions

Categories: PCR, Polymerases & Amplification Technologies, Phosphatase & Sulfurylase Products, Nuclease Products

Application: Rapid PCR Cleanup Enzyme Set degrades excess primers and dNTPs for purification of PCR products prior to sequencing.
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Description Advantages and Features
Properties and Usage Quality Control

Description

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The NEB Rapid PCR Cleanup Enzyme Set consists of two recombinant enzymes, Exonuclease I (Exo I) and Shrimp Alkaline Phosphatase (rSAP). It allows for rapid and complete enzymatic degradation of residual PCR primers and dephosphorylation of dNTPs subsequent to PCR. The Rapid PCR Cleanup Enzyme Set enables direct downstream sequencing without the need for additional purification steps. The Rapid PCR Cleanup Enzyme Set is added directly to the PCR reaction after thermal cycling and is 100% compatible with commonly used PCR reaction buffers.

Product Source
rSAP: A Pichia pastoris strain that carries the shrimp alkaline phosphatase gene from Northern shrimp Pandalus borealis (1,2).
Exonuclease I: An Escherichia coli strain that carries an extra gene construct coding for E. coli Exonuclease I.

asap fig1_gel_white

Figure 1. Agarose gel showing three different PCR products before and after Rapid PCR Cleanup Enzyme Set treatment. No loss of PCR product was detected, irrespective of amplicon size.
asap-fig1-1

Figure 2. PCR cleanup is important prior to sequencing. Panel A shows a sample that has undergone Rapid PCR Cleanup Enzyme Set treatment, while panel B shows a sample without this treatment. Rapid PCR Cleanup Enzyme Set treatment resulted in significant improvement in overall sequence quality. 

 

Reagents supplied
The following reagents are supplied with this product:

  Store at (°C)
Exonuclease I (#M0561) -20
rSAP (#M0562) -20

Advantages and Features

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  • Fast 15 minute protocol
  • Add directly to PCR product
  • 100% Sample Recovery
  • Scalable for different reaction sizes
  • No interference on downstream applications
  • Easy to automate

Properties and Usage

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Reaction Conditions
Incubate at 37°C for 5 minutes

Heat Inactivation
80°C for 10 minutes

Storage Temperature
-20°C

Quality Control

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Quality Control Assays
The following Quality Control Tests are performed on each new lot and meet the specifications designated for the product. Individual lot data can be found on the Product Summary Sheet/Datacard or Manual which can be found in the Supporting Documents section of this page.

Functional Assay
Functionally tested using a PCR product of the malE gene generated from 50 ng of pMALc-5X plasmid template in 30 PCR cycles. The amplicons were sequenced by Applied Biosystems® 3130xl Genetic Ananlyzer using Big Dye Terminator chemistry. The PCR product sequences were readable for 600 bp by Lasergene® software analysis at QV20+ (Phred20).

References

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1. Olsen, R.L. et al. (1991). Comp. Biochem. Physiol. 99B(4):, 755-761.
2. Nilsen, I.W. et al. (2001). Comp. Biochem. Physiol. 129B(4):, 853-861.

Legal and Disclaimers

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Applied Biosystems® is a registered trademark of Applera Corporation or subsidiaries of Applera Corporation in the U.S. and certain other countries.

Lasergene® is a registered trademark of DNAStar, Inc.

Supporting Documents

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Material Safety Datasheets The following is a list of Material Safety Data Sheets (MSDS) that apply to this product to help you use it safely. The following file naming structure is used to name these document files: [Product Name] MSDS. For international versions please contact us at info.de@neb.com.

Exonuclease I MSDS

Shrimp Alkaline Phosphatase (rSAP) MSDS

Datacards The Product Summary Sheet, or Data Card, includes details for how to use the product, as well as details of its formulation and quality controls. The following file naming structure is used to name the majority of these document files: [Catalog Number]Datasheet-Lot[Lot Number]. For those product lots not listed below, please contact NEB at info.de@neb.com for appropriate document. For additional technical information, please contact us at techsupport.de@neb.com.

E2622Datasheet-Lot0011411