One buffer for 176 enzymes. It's that easy.

One buffer for 176 enzymes. It's that easy.

Thermo Scientific FastDigest restriction enzymes are an advanced line of enzymes which offer:

  • Complete digestion in 5-15 minutes
  • 100% buffer compatibility with downstream applications
  • Direct loading on gels

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Supported by over 40 years of experience

Through the FastDigest collection, you have access to one of the largest collections of restriction enzymes in the industry. Decades of expertise in enzymology, the large selection of enzyme isoschizomers, and the ability to produce enzymes of exceptional purity from our state-of-the-art manufacturing facilities helped enable the creation of the first simplified restriction enzyme system using a universal buffer.

Advantages of FastDigest restriction enzymes

  FastDigest restriction enzymes Conventional restriction enzymes
Buffer system One Universal Buffer Up to 20 Buffers
Double/multiple restriction digestion Unlimited – all 176 enzymes are 100% active in one buffer Limited by buffer compatibility
Reaction Time 5–15 minutes 1 hour–overnight
Direct loading on gels Yes No
Downstream applications 100% buffer compatibility Partial compatibility
Star activity No Yes
Activity definintion 1 µL of FastDigest enzyme cleaves1 µg of substrate DNA in 5 to 15 minutes in FastDigest buffer 1 unit of enzyme hydrolyzes 1µg of lambda DNA in 60 minutes in an optimal buffer for an enzyme

100% activity in a single buffer

The FastDigest Green Buffer and Thermo Scientific FastDigest Buffer are proprietary digestion buffers which support 100% activity of all FastDigest restriction enzymes. This system allows for double and multiple digestions with any combination of enzymes. No sequential digestions or buffer changes are needed.

Five minute single, double and triple digestions in FastDigest Green Buffer

Five minute single, double and triple digestions in FastDigest Green Buffer

Five minute plasmid DNA digestions with:

  • Lane M: Thermo Scientific GeneRuler Express Ladder (Cat. No. SM1553)
  • Lane 1: Control, undigested plasmid DNA
  • Lane 2: FastDigest EcoRI
  • Lane 3: FastDigest EcoRI and FastDigest KpnI
  • Lane 4: FastDigest EcoRI, FastDigest KpnI and FastDigest SmaI

100% buffer compatibility with downstream applications

DNA/RNA modifying enzymes, such as ligases, phosphatases, kinases and mesophilic DNA polymerases have 100% activity in FastDigest and FastDigest Green Buffer. Therefore, enzymes used in downstream applications can be directly added to the FastDigest reaction mix. No buffer changes or purification steps are needed.

DNA/RNA Modifying Enzyme Activity in FastDigest Green Buffer/FastDigest Buffer
DNA Polymerase I, E. coli 100%
Klenow Fragment 100%
Klenow Fragment, exo- 100%
T4 DNA Polymerase 100%
T7 DNA Polymerase 100%
T4 DNA Ligase* 75–100%
FastAP Thermosensitive Alkaline Phosphatase 100%
T4 Polynucleotide Kinase 100%

* 0.5 mM ATP is required for T4 DNA Ligase activity.


Complete digestion in 5-15 minutes

Double and multiple digestions in one buffer in 5-15 minutes:

  • Saves time and effort, increasing throughput.
  • Digestion times are provided for all types of DNA templates (plasmid DNA, PCR product, genomic DNA).
  • No overnight digestions are required for any template.
  • Star activity is eliminated due to short reaction times.
Conventional restriction enzymes
Sequential digestion
FastDigest restriction enzymes
One reaction mixture
Reaction setup for Apal ~2 min Reaction set up with FastDigest ApaI and XhoI ~2 min
Incubation 60 min Incubation 5 min
Reaction setup for Xhol ~2 min
Incubation 60 min
TOTAL TIME >2 hours TOTAL TIME  7 min

Direct loading on gels

Five minute single, double and triple digestions in FastDigest Green Buffer

As an added convenience we developed the FastDigest Green Buffer which offers the same performance as the colorless FastDigest Buffer but enables direct loading of the reaction mixture on gels. The 10X FastDigest Green Buffer includes a density reagent and two tracking dyes for direct loading. The blue dye migrates with 3–5 kb DNA fragments in a 1% agarose gel and has an excitation peak of 424 nm. The yellow dye migrates faster than 10 bp DNA fragments in a 1% agarose gel and has an excitation peak of 615 nm.

FastDigest Green Buffer

Reaction mixture containing FastDigest Green Buffer:

  • Lane 1: Loaded into gel well, before electrophoresis
  • Lane 2: Separated, after electrophoresis

Short protocol times with no star activity

FastDigest enzymes in combination with FastDigest (Green) Buffer are designed to eliminate star activity:

  • Short incubation time (5–15 min), no need for prolonged digestions.
  • Universal buffer with optimal pH and salt concentration for all enzymes.
  • Optimal enzyme and glycerol concentrations.

Conventional restriction enzymes may display star or "relaxed" activity due to prolonged incubation times, high enzyme and/or glycerol concentration, high pH values or low ionic strength. By addressing all these issues, FastDigest enzymes enable single, double and even triple digestion in 5 minutes without any signs of star activity.

Five minute single, double and triple digestions in FastDigest Green Buffer

Five minute single and double digestion of plasmid DNA with no signs of star activity

Five minute plasmid DNA digestions with:

  • Lane 1: FastDigest SmaI
  • Lane 2: FastDigest EcoRI
  • Lane 3: FastDigest SmaI and FastDigest EcoRI double digest
  • Lane M: GeneRuler 1 kb Plus DNA Ladder (Cat. No. SM1333)
  • Lane C: Control, Undigested plasmid DNA