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Kpn2I (BspEI)

5'...T↓C C G G A...3'
3'...A G G C C↑T...5'
Available as a FastDigest enzyme for rapid DNA digestion

FastDigest enzyme available Tango buffer for 100% activity Optimal incubation at 55°C Cleavage blocked or impaired by CpG methylation Thermal inactivation at 80°C in 20 min Genome qualified Recombinant enzyme LO certified

The Kpn2I (BspEI) restriction enzyme recognizes T^CCGGA sites and cuts best at 55°C in Tango buffer (Isoschizomers: AccIII, Aor13HI, BseAI, Bsp13I, BspEI, MroI)
  
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Lambda DNA digested with Kpn2I (BspEI)

Lambda DNA digested with Kpn2I (BspEI), 0.7% agarose, 24 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.

Highlights

  • 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

Applications

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

Note

For methylation sensitivity refer to product specifications.

  
Storage Condition-20 C
HazardousNo
Compatible EndsBshTI, BsaWI, Cfr9I, Cfr10I, Eco88I, NgoMIV, SgrAI.
Conditions for 100% Activity
  • X Buffer Tango: 33 mM Tris-acetate (pH 7.9 at 37°C), 10 mM Mg-acetate, 66 mM K-acetate, and 0.1 mg/mL BSA
  • Incubate at 55°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.
NoteIncubation at 37°C results in 50% activity.
Storage BufferKpn2I is supplied in: 10 mM Tris-HCl (pH 7.4 at 25°C), 100 mM KCl, 1 mM DTT, 1 mM EDTA, 0.2 mg/mL BSA, and 50% (v/v) glycerol.

Reaction conditions

Recommended buffer for 100% activity Optimal temperature Enzyme activity in Thermo Scientific buffers, % Tango buffer for double digestion
B (blue)
1X		 G (green)
1X		 O (orange)
1X		 R (red)
1X		 Tango
(yellow)
1X / 2X
Tango 55°C 50-100 50-100 0-20 20-50 100 50-100 1X or 2X

Methylation Effects

Methylation type Sequence Cleavage effect
Dam(GATC) 5'...TCCGGm6A TC...3'
3'...AGGCC Tm6AG...5'		 No effect
Dam(GATC) 5'...Gm6A TCCGGm6A TC...3'
3'...C Tm6AGGCC Tm6AG...5'		 Not determined
CpGTCCGGA Blocked

Cleavage efficiency close to the termini of PCR fragments

bp from the recognition site to fragment end
1 2 3 4 5
0 50-100

Number of recognition sites in DNA molecules

Lambda ΦX174 M13mp18/19 pBR322 puc18/19 pUC57
24 0 0 1 0 0
pTZ19R/U pTZ57R pBluescriptIIKS(-/+) pBluescriptIISK(-/+) pACYC177 pACYC184
0 0 0 0 0 2

New sites generated by ligation

Newly generated recognition sites resulting from ligation of protruding compatible DNA ends

Recognition Sequence Second Enzyme Enzymes recognizing newly generated recognition sequence
T^CCGGA
  • BsaWI* (W^CCGGA)
  • BsaWI
  • HpaII/FastDigest HpaII
  • Hpy188III
  • Kpn2I (BspEI)/FastDigest Kpn2I
  • MspI (HpaII)/FastDigest MspI
  • BsaWI* (W^CCGGT)
  • BshTI (AgeI)/FastDigest AgeI (BshTI) (A^CCGGT)
  • Cfr10I (BsrFI)/FastDigest BsrFI (Cfr10I)* (R^CCGGT)
  • SgrAI* (CR^CCGGTG)
  • BsaWI
  • HpaII/FastDigest HpaII
  • MspI (HpaII)/FastDigest MspI
  • Cfr10I (BsrFI)/FastDigest BsrFI (Cfr10I)* (R^CCGGC)
  • MreI (Sse232I)/FastDigest MreI (CG^CCGGCG)
  • NgoMIV (G^CCGGC)
  • SgrAI* (CR^CCGGCG)
  • HpaII/FastDigest HpaII
  • MspI (HpaII)/FastDigest MspI
  • Cfr9I (XmaI) (C^CCGGG)
  • Eco88I (AvaI)/FastDigest AvaI (Eco88I)* (C^CCGGG)
  • BcnI (NciI)/FastDigest NciI (BcnI)
  • Bme1390I (ScrFI)/FastDigest ScrFI (Bme1390I)
  • BssKI
  • HpaII/FastDigest HpaII
  • MspI (HpaII)/FastDigest MspI

Newly generated recognition sites resulting from fill-in of 5'-overhang and self-ligation

Recognition Sequence Newly generated sequence after reaction Enzymes that cleave the newly generated sequence
T^CCGGATCCGGCCGGA
  • Bsh1285I (BsiEI)/FastDigest BsiEI (Bsh1285I)
  • BsuRI (HaeIII)/FastDigest HaeIII (BsuRI)
  • EaeI
  • CviJI
  • Eco52I (EagI)/FastDigest EagI (Eco52I)
  • [HpaII/FastDigest HpaII]
  • [MspI (HpaII)/FastDigest MspI]