KpnI, HC (50 U/l)

Q: What are key factors promoting star activity?

Star activty may be contributed by:• Prolonged incubation• High enzyme concentration• High glycerol concentration (usually 5% or higher)• Small reaction volume Find additional tips, troubleshooting help, and resources within our Restriction Enzyme Cloning Support Center .

5' G G T A C ↓C 3' 3' C ↑C A T G G 5' Das Thermo Scientific RestriktionsenzymWeitere Informationen

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Katalognummer	Menge
ER0521	4000 E
ER0522	5 x 4000 E

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Katalognummer ER0521

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4000 E

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5' G G T A C ↓ C 3'
3' C ↑ C A T G G 5'

Das Thermo Scientific Restriktionsenzym KpnI erkennt GGTAC^C-Stellen und schneidet am besten bei 37°C in seinem eigenen einzigartigen Puffer. Unter Reaktionsbedingungen für Restriktionsenzym finden Sie eine Tabelle mit Angaben zu Enzymaktivität, Bedingungen für doppelte Verdauung und Hitzeinaktivierung für diese und andere Restriktionsenzyme. Isoschizomere: Asp718I.

Herkömmliche Thermo Scientific Restriktionsendonukleasen sind eine große Ansammlung hochwertiger Restriktionsenzyme, die für den Einsatz in einem der Puffer des Fünf-Puffer-Systems optimiert sind. Darüber hinaus bietet der universelle Tango Puffer Vorteile bei Doppelverdauungen. Alle Enzyme zeigen unter den empfohlenen Puffer- und Reaktionsbedingungen 100 % Aktivität. Zur Gewährleistung gleichbleibender Leistungsfähigkeit enthalten Thermo Scientific Restriktionsenzym-Puffer vorgemischtes BSA, welches die Stabilität zahlreicher Enzyme verbessert und Schadstoffe bindet, die in DNA-Präparaten enthalten sein können.

Eigenschaften

• überragende Qualität — strenge Qualitätskontrolle und branchenführender Fertigungsprozess
• praktisches, farbcodiertes Fünf-Puffer-System
• mit universellem Tango Puffer für Doppelverdauungen
• BSA in Reaktionspuffern vorgemischt
• große Auswahl an Restriktionsendonuklease-Spezifikationen

Anwendungen

• molekulare Klonierung
• Kartierung von Restriktionsstellen
• Genotypisierung
• Southern Blotting
• Restriktionsfragmentlängenpolymorphismus (RFLP)
• SNP

Hinweis: Angaben zur Methylierungssensitivität finden Sie in den Produktspezifikationen.

Nur für Forschungszwecke. Nicht zur Verwendung bei diagnostischen Verfahren.

Specifications

Kompatibler PufferSpezial-Puffer (10x Puffer KpnI)

ProdukttypRestriktionsenzym

Menge4000 E

Konzentration10 U/μl

EnzymKpn I

MethylierungsempfindlichkeitNicht empfindlich für Dam-Methylierung, Dcm-Methylierung und CpG-Methylierung, Dcm-Methylierung und CpG-Methylierung, Not CpG Methylation-Sensitive

Optimale Reaktionstemperatur37°C

ForschungskategorieHerkömmliches Klonen

Empfindlich gegen HitzeinaktivierungJa

Typ IIS RENein

Unit SizeEach

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Häufig gestellte Fragen (FAQ)

Can I double digest my DNA using a Thermo Scientific conventional restriction enzyme and a FastDigest restriction enzyme?

For optimal results with fast reaction and 100% buffer compatibility, we highly recommend using FastDigest restriction enzymes in double digestion. In certain cases however, it may be possible to perform double digestion using a mix of Thermo Scientific conventional and Fastdigest restriction enzymes. For specific recommendations, please contact our technical service with detailed information about the enzymes and DNA template you plan to use.

Why do you recommend only 2 µL of 10X Reaction Buffer when digesting unpurified PCR product in a 30 µL reaction?

We recommend only 2 µl 10X Buffer in digestion of unpurified PCR products in 30 ul since salts and ions from the PCR reaction would be carried over to the digestion reaction.

Find additional tips, troubleshooting help, and resources within our Restriction Enzyme Cloning Support Center.

What are key factors promoting star activity?

Star activty may be contributed by:

• Prolonged incubation
• High enzyme concentration
• High glycerol concentration (usually 5% or higher)
• Small reaction volume

Find additional tips, troubleshooting help, and resources within our Restriction Enzyme Cloning Support Center.

Unexpected DNA bands were observed on agarose gel electrophoresis after restriction digestion. What may have caused this?

Unexpected cleavage patterns may be caused by the following reasons:

• Star activity of the restriction enzyme: Make sure to follow the reaction recommendations as specified in the protocol. Star activity may be improved by changing several key factors such as decreasing the reaction time, increasing the reaction volume, and decreasing the enzyme amount.

• Partial or incomplete cleavage (incomplete restriction reaction): Efficiency of the enzyme can be improved by adding more enzyme, prolonging the reaction time, or purifying DNA samples to remove inhibitory contaminants.

• Contamination with non-specific endonucleases: Non-specific endonucleases may be introduced to the DNA sample and/or the enzyme from improper handling, pipetting, etc.

•Improper reaction setup: Mix the digestion reaction thoroughly.

Find additional tips, troubleshooting help, and resources within ourRestriction Enzyme Cloning Support Center.

What are possible reasons for incomplete/failed restriction digestion?

The main reason for DNA cleavage reaction failure is the presence of contaminating inhibitors in the template DNA (for example: phenol, chloroform, detergents, ethanol, excess salts, EDTA, etc.). The best way to troubleshoot is to perform control reactions:

1) negative control (experimental DNA in the reaction buffer without the restriction enzyme) to access degradation of DNA by contaminants in the DNA template and/or reaction buffer
2) positive control reaction I (digestion of highly pure control DNA with the restriction enzyme) to access reaction conditions and enzyme activity
3) positive control reaction II (highly pure control DNA + experimental DNA + Restriction Enzyme) to access possible issues with the experimental DNA.

In addition, please check for sensitivity of the restriction enzymes to template DNA methylation.

Find additional tips, troubleshooting help, and resources within our Restriction Enzyme Cloning Support Center.

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