KpnI

Now comes supplied with 1 vial of Gel Loading Dye, Purple (6X), (B7024). KpnI has an HF version supplied with CutSmart Buffer.
recombinant $name $name incubation temp heat inactivation no blue white
Kpn-I-cutsite_1_v1_000021
Catalog #SizeConcentrationPriceQtyAdd to Cart
R0142S4,000 units10,000 units/ml$63.00Add to Cart
R0142L20,000 units10,000 units/ml$254.00Add to Cart
R0142M20,000 units50,000 units/ml$254.00Add to Cart
  
Categories:
Restriction Endonucleases: H-M,
Time-Saver™ Qualified Restriction Enzymes
Applications:
Restriction Enzyme Digestion

Description

KpnI has a High Fidelity version KpnI-HF™ (NEB #R3142).

High Fidelity (HF ™) Restriction Enzymes have 100% activity in CutSmart ™ Buffer; single-buffer simplicity means more straightforward and streamlined sample processing. HF enzymes also exhibit dramatically reduced star activity. HF enzymes are all Time-Saver qualified and can therefore cut substrate DNA in 5-15 minutes with the flexibility to digest overnight without degradation to DNA. Engineered with performance in mind, HF restriction enzymes are fully active under a broader range of conditions, minimizing off-target products, while offering flexibility in experimental design.

Product Source

An E. coli strain that carries the KpnI gene from Klebsiella pneumoniae OK8 (ATCC 49790).

Reagents Supplied

The following reagents are supplied with this product:

Store at (°C)Concentration
NEBuffer 1.1-2010X

Properties and Usage

Unit Definition

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

Reaction Conditions

1X NEBuffer 1.1
Incubate at 37°C

1X NEBuffer 1.1:
10 mM Bis-Tris-Propane-HCl
10 mM MgCl2
100 μg/ml BSA
pH 7 @ 25°C

Activity in NEBuffers

NEBuffer 1.1: 100%
NEBuffer 2.1: 75%
NEBuffer 3.1: 10%
CutSmart® Buffer: 50%

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: Not Sensitive
dcm methylation: Not Sensitive
CpG Methylation: Not Sensitive

Unit Assay Conditions

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

Notes

  1. Acc65I is an isoschizomer of KpnI.
  2. KpnI produces a 4-base 3´extension, whereas Acc65I produces a 4-base 5´ extension.
  3. May exhibit Star Activity in CutSmart Buffer.
  4. 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.
  5. Star activity may result from extended digestion, high enzyme concentration or a glycerol concentration of >5%

FAQs

  1. How can this enzyme be inactivated?
  2. My enzyme is no longer Time-Saver™ qualified. What happened?
  3. What effect does BSA have on the performance of NEB’s restriction enzymes when included in the new buffers?
  4. How can I access the old NEBuffer Activity Chart?
  5. Why is my Restriction Enzyme not cutting DNA?
  6. Why do I see a DNA smear on an agarose gel after a restriction digest?
  7. Why do I see additional DNA bands on my gel after a restriction digest?
  8. How many nucleotides do I have to add adjacent to the RE recognition site in order to get efficient cutting?

Protocols

  1. Optimizing Restriction Endonuclease Reactions
  2. Double Digest Protocol with Standard Restriction Enzymes
  3. Time-Saver Protocol for Restriction Enzyme Digests

Selection Charts

Usage Guidelines & Tips

Troubleshooting Guides

Interactive Tools

Quality Control

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. Further information regarding NEB product quality can be found here.
  • Blue-White Screening (Terminal Integrity):
    A sample of DNA vector linearized with a 10-fold excess of a restriction endonuclease, religated and transformed into an E. coli strain expressing the LacZ beta fragment gene results in less than 1% white colonies.
  • Endonuclease Activity (Nicking, Buffer):
    The buffer is tested in a reaction containing a supercoiled DNA substrate. After incubation for 4 hours the percent converted to the nicked form is determined by agarose gel electrophoresis.
  • Exonuclease Activity (Radioactivity Release):
    The product is tested in a reaction containing a radiolabeled mixture of single and double-stranded DNA. After incubation for 4 hours the exonuclease activity is determined by the % release of radioactive nucleotides.
  • Ligation and Recutting (Terminal Integrity):
    After an over-digestion of DNA with a restriction endonuclease the percentage of the DNA fragments ligated with T4 DNA ligase and the percentage that can be recut are determined by agarose gel electrophoresis.
  • Non-Specific DNase Activity (16 hour, DNA):
    The DNA is tested in a reaction under standard reaction conditions. After incubation for 16 hours there is no detectable degradation of the DNA as determined by agarose gel electrophoresis.

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]

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]

Safety Data Sheet

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