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FastDigest DpnI

5'...G m6A↓  T C...3'
3'...C   T↑m6A G...5'

FastDigest buffer for 100% activity Optimal incubation at 37°C Thermal inactivation at 80°C in 5 min Recombinant enzyme LO certified

The FastDigest DpnI restriction enzyme recognizes Gm6A^TC sites and cuts best at 37°C in 5-15 minutes using universal Yellow Tango Buffer (Isoschizomers: MalI)
  
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Lambda DNA digested with DpnI

Lambda DNA digested with DpnI, 1.4% agarose, 22 cleavage sites

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
Formulation1 μL of enzyme (1 FDU) cleaves 1 μg of pBR322 DNA (dam methylated) 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 DpnI is formulated to digest up to:

  • 1 µg of pBR322 DNA (dam methylated) in 5 minutes
  • 1 µg of plasmid DNA in 5 minutes
  • Digestion of PCR products is not applicable
  • 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

(pBR322 DNA)		 5 not applicable 10 1 80°C, 5 min 16

Methylation Effects

Methylation type Sequence Cleavage effect
CpG5'...Gm6A Tm5C G...3'

3'...C Tm6A Gm5C...5'		 Cleaves only Dam methylated DNA
Dam(GATC)Gm6ATC Cleaves only Dam methylated DNA

Number of recognition sites in DNA molecules

Lambda ΦX174 M13mp18/19 pBR322 puc18/19 pUC57
116 0 7 22 15 15
pTZ19R/U pTZ57R pBluescriptIIKS(±) pBluescriptIISK(±) pACYC177 pACYC184
15 15 15 15 22 15

New sites generated by ligation

Newly generated recognition sites resulting from ligation of blunt DNA ends

Recognition sequence Second restriction enzyme Restriction enzymes cleaving the newly generated recognition sequence
GA^TC
  • AjiI (BmgBI)* (CAC^GTC)
  • ZraI (GAC^GTC)
  • HinfI/FastDigest HinfI
  • PleI
  • SchI (MlyI)/FastDigest MlyI (SchI)
  • Ecl136II (EcoICRI)/FastDigest Ecl136II (GAG^CTC)
  • MbiI (BsrBI)/FastDigest BsrBI (MbiI)* (CCG^CTC)
  • HinfI/FastDigest HinfI
  • Eco147I (StuI)/FastDigest StuI (Eco147I) (AGG^CCT)
  • SetI
  • Eco32I (EcoRV)/FastDigest EcoRV (Eco32I) (GAT^ATC)
  • HinfI/FastDigest HinfI
  • PfeI (TfiI)/FastDigest TfiI (PfeI)
  • Eco47III (AfeI)/FastDigest AfeI (Eco47III) (AGC^GCT)
  • AluI/FastDigest AluI
  • CviJI
  • SetI
  • EheI (SfoI)/FastDigest EheI (GGC^GCC)
  • CviJI
  • FspAI/FastDigest FspAI* (RTGC^GCAC)
  • Alw21I (BsiHKAI)/FastDigest Alw21I
  • SduI (Bsp1286I)/FastDigest Bsp1286I (SduI)
  • SspI/FastDigest SspI (AAT^ATT)
  • TasI (Tsp509I)/FastDigest Tsp509I (TasI)