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5'...G G T A CC...3'
3'...CC A T G G...5

Available as a FastDigest enzyme for rapid DNA digestion

FastDigest enzyme available Unique buffer for 100% activity Optimal incubation at 37°C Thermal inactivation at 80°C in 20 min Genome qualified High concentration available LO certified

The KpnI restriction enzyme recognizes GGTAC^C sites and cuts best at 37°C in its own unique buffer (Isoschizomers: Asp718I)
Lambda DNA digested with KpnI

Lambda DNA digested with KpnI, 0.7% agarose, 2 cleavage sites

Thermo Scientific conventional restriction endonucleases are a large collection of high quality restriction enzymes, optimized to work in one of the buffers of the Five Buffer System. In addition, the universal Tango buffer is provided for convenience in double digestions. All of the enzymes exhibit 100% activity in the recommended buffer and reaction conditions. To ensure consistent performance, Thermo Scientific restriction enzyme reaction buffers contain premixed BSA, which enhances the stability of many enzymes and binds contaminants that may be present in DNA preparations.


  • Superior quality – stringent quality control and industry leading manufacturing process
  • Convenient color-coded Five Buffer System
  • Includes universal Tango buffer for double-digestions
  • BSA premixed in reaction buffers
  • Wide selection of restriction endonuclease specificities


  • Molecular cloning
  • Restriction site mapping
  • Genotyping
  • Southern Blot
  • Restriction fragment length polymorphism (RFLP)
  • SNP


For methylation sensitivity refer to product specifications.

Conditions for 100% Activity
  • 1X Buffer KpnI: 10 mM Tris-HCl (pH 7.5 at 37°C.C), 10 mM MgCl2, 0.02% Triton X-100, and 0.1 mg/mL BSA
  • Incubate at 37°C
Digestion of Agarose-embedded DNAMinimum 5 units of the enzyme are required for complete digestion of 1 µg of agarose-embedded lambda DNA in 16 hours.
Double DigestionPerform double digestion using DoubleDigest.
IsoschizomersSearch for commercial isoschizomers using REsearch.
NoteHigh glycerol (> 5%) concentrations, pH > 8.0 or a large excess of enzyme may result in star activity.
Storage BufferKpnI is supplied in: 10 mM Tris-HCl (pH 7.5 at 25°C), 50 mM KCl, 1 mM DTT, 0.1 mM EDTA, 0.2 mg/mL BSA and 50% (v/v) glycerol.
Storage Condition-20 C

Reaction conditions

Recommended buffer for 100% activity Optimal temperature Enzyme activity in Thermo Scientific buffers, % Tango buffer for double digestion
B (blue)
G (green)
O (orange)
R (red)
1X / 2X
KpnI Buffer [Unique] 37°C 20-50 0-20 0-20 0-20 20-50 0-20 1X

Methylation Effects

Methylation type Sequence Cleavage effect
Dcm(CCWGG)5'...GGTACm5CW GG...3'
3'...CCATG GWm5CC...5'
No effect
Dcm(CCWGG)5'...Cm5CW GGTACm5CW GG...3'
3'...G GWm5CCATG GWm5CC...5'
No effect
CpG5'...m5C GGTACm5C G...3'
3'...Gm5CCATG Gm5C...5'
No effect

Cleavage efficiency close to the termini of PCR fragments

bp from the recognition site to fragment end
1 2 3 4 5
50 to 100

Number of recognition sites in DNA molecules

Lambda ΦX174 M13mp18/19 pBR322 puc18/19 pUC57
2 0 1 0 1 1
pTZ19R/U pTZ57R pBluescriptIIKS(-/+) pBluescriptIISK(-/+) pACYC177 pACYC184
1 1 1 1 0 0