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FastDigest XbaI

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

FastDigest buffer for 100% activity Optimal incubation at 37°C Cleavage blocked or impaired by overlapping Dam methylation Thermal inactivation at 65°C in 20 min LO certified

The FastDigest XbaI restriction enzyme recognizes T^CTAGA sites and cuts best at 37°C in 5-15 minutes using universal Yellow Tango Buffer
  
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Lambda DNA digested with XapI

Lambda DNA (dam-), 0.7% agarose, 1 cleavage site

Thermo Scientific FastDigest enzymes are an advanced line of fast restriction enzymes that are all 100% active in the universal FastDigest and FastDigest Green reaction buffers.

The universal buffer allows rapid single-, double- or multiple DNA digestion within 5-15 minutes eliminating any need for buffer change or subsequent DNA clean-up steps. DNA modifying enzymes, such as Klenow Fragment, T4 DNA Ligase, alkaline phosphatases and T4 DNA Polymerase all have 100% activity in FastDigest Buffer. Therefore, enzymes for downstream applications can be directly added to the FastDigest reaction mix.

For additional convenience FastDigest Green Buffer includes a density reagent and two tracking dyes for direct loading of digestion reaction products on gels.

Short incubation times and optimal composition of the universal FastDigest Buffer eliminate star activity effects.

Features

  • 100% activity of all FastDigest enzymes in the universal buffer
  • 100% buffer compatibility with downstream applications
  • Complete digestion in 5-15 minutes
  • Direct loading on gels
  • No star activity
  • 176 FastDigest enzymes available

Applications

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

Note

The FastDigest Green Buffer offers the same high performance in DNA digestion and downstream applications as the colorless FastDigest Buffer. For applications that require product analysis by fluorescence excitation (e.g. concentration measurements in UV light) the colorless FastDigest Buffer is recommended.

For methylation sensitivity refer to product specifications.

  
Storage Condition-20 C
HazardousNo
Compatible EndsFastDigest AvrII (XmaJI), FastDigest NheI, FastDigest SpeI (BcuI), FastDigest StyI (Eco130I), AvrII, BcuI, Eco130I, NheI, SpeI, StyI, XmaJI.
Formulation1 µL of enzyme (1 FDU) cleaves 1 μg of lambda DNA dam-/SmaI fragments in 5 minutes at 37°C in 1X FastDigest Buffer.
IsoschizomersSearch for commercial isoschizomers using REsearch.
Recommended Reaction Conditions

1X FastDigest buffer or 1X FastDigest Green buffer at 37°C.
1 µL of FastDigest XbaI is formulated to digest up to:

  • 1 µg of lambda DNA dam- /SmaI fragments in 5 minutes
  • 1 µg of plasmid DNA in 5 minutes
  • 0.2 µg of PCR product in 5 minutes
  • 1 µg of genomic DNA in 10 minutes or 5 µg of genomic DNA in 60 minutes

Reaction conditions

Reaction temperature Digestion time with 1 µL of FastDigest enzyme, min bp from end of DNA required for complete digestion Thermal inactivation Incubation time without star activity, hours
Lambda, 1 µg/20 µL Plasmid DNA, 1 µg/20 µL PCR product, ~0.2 µg/30 µL Genomic DNA, 1 µg/10 µL
37°C 5 5 5 10 2 65°C, 20 min 16

Methylation Effects

  • Dam: may overlap - blocked.
  • Dcm: never overlaps - no effect.
  • CpG: never overlaps - no effect.
  • EcoKI: never overlaps - no effect.
  • EcoBI: never overlaps - no effect.
Methylation type Sequence Cleavage effect
Dam(GATC)5'...TCTAGm6A TC...3'
3'...AGATC Tm6AG...5'		 Blocked

Number of recognition sites in DNA molecules

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

New sites generated by ligation

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

Recognition SequenceSecond EnzymeEnzymes recognizing newly generated recognition sequence
T^CTAGA
  • BcuI (SpeI)/FastDigest SpeI (BcuI) (A^CTAGT)
  • Eco130I (StyI)/FastDigest StyI (Eco130I)* (C^CTAGG)
  • NheI/FastDigest NheI (G^CTAGC)
  • XmaJI (AvrII)/FastDigest AvrII (XmaJI) (C^CTAGG)
  • FspBI (BfaI)/FastDigest BfaI (FspBI)

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

Recognition SequenceNewly generated sequence after reactionEnzymes that cleave the newly generated sequence
T^CTAGATCTAGCTAGA
  • AluI/FastDigest AluI
  • CviJI
  • [FspBI (BfaI)/FastDigest BfaI (FspBI)]
  • SetI