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Home | Molecular Biology | Restriction & Modifying Enzymes | Restriction Enzymes | Conventional

Eco47III (AfeI)

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

FastDigest enzyme available O buffer for 100% activity Optimal incubation at 37°C Cleavage blocked or impaired by CpG methylation Thermal inactivation at 80°C in 20 min Genome qualified Recombinant enzyme LO certified

The Eco47III (AfeI) restriction enzyme recognizes AGC^GCT sites and cuts best at 37°C in O buffer (Isoschizomers: AfeI, Aor51HI)
  
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Lambda DNA digested with Eco47III (AfeI)

Lambda DNA digested with Eco47III (AfeI), 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.

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
Conditions for 100% Activity
  • 1X Buffer O: 50 mM Tris-HCl (pH 7.5 at 37°C), 10 mM MgCl2, 100 mM NaCl, 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.
Storage BufferEco47III is supplied in: 10 mM Tris-HCl (pH 7.4 at 25°C), 100 mM KCl, 1 mM DTT, 1mM 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
O (Orange) 37°C 0-20 20-50 100 100 50-100 100 1X or 2X

Methylation Effects

  • Dam: never overlaps - no effect.
  • Dcm: never overlaps - no effect.
  • CpG: completely overlaps - blocked.
  • EcoKI: never overlaps - no effect.
  • EcoBI: may overlap - effect not determined.
Methylation type Sequence Cleavage effect
CpGAGCGCT Blocked

Cleavage efficiency close to the termini of PCR fragments

bp from the recognition site to fragment end
1 2 3 4 5
0 0-20 50-100

Number of recognition sites in DNA molecules

Lambda ΦX174 M13mp18/19 pBR322 puc18/19 pUC57
2 0 2 4 0 0
pTZ19R/U pTZ57R pBluescriptIIKS(-/+) pBluescriptIISK(-/+) pACYC177 pACYC184
0 0 0 0 2 5

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
AGC^GCT
  • AanI (PsiI)/FastDigest PsiI (AanI) (TTA^TAA)
  • Bst1107I (BstZ17I)/FastDigest BstZ17I (Bst1107I) (GTA^TAC)
  • DpnI/FastDigest DpnI (GA^TC)
  • FaiI* (YA^TA)
  • FaiI* (YA^TG)
  • AluI/FastDigest AluI
  • CviJI
  • SetI
  • AluI/FastDigest AluI (AG^CT)
  • Bsh1236I (BstUI)/FastDigest Bsh1236I (CG^CG)
  • Bsp68I (NruI)/FastDigest NruI (RruI) (TCG^CGA)
  • BsuRI (HaeIII)/FastDigest HaeIII (BsuRI) (GG^CC)
  • CviJI* (RG^CC)
  • CviJI* (RG^CT)
  • Eco147I (StuI)/FastDigest StuI (Eco147I) (AGG^CCT)
  • HpyCH4V (TG^CA)
  • MlsI (MscI)/FastDigest MscI (MlsI) (TGG^CCA)
  • MspA1I* (CMG^CTG)
  • PvuII/FastDigest PvuII (CAG^CTG)
  • CviJI
  • Ecl136II (EcoICRI)/FastDigest Ecl136II (GAG^CTC)
  • MbiI (BsrBI)/FastDigest BsrBI (MbiI)* (CCG^CTC)
  • CviJI
  • MnlI/FastDigest MnlI
  • Eco32I (EcoRV)/FastDigest EcoRV (Eco32I) (GAT^ATC)
  • LweI (SfaNI)/FastDigest SfaNI (BmsI)
  • EheI (SfoI)/FastDigest EheI (GGC^GCC)
  • FastDigest HaeII (BfoI)
  • HhaI/FastDigest HhaI
  • Hin6I (HinP1I)/FastDigest HinP1I (Hin6I)
  • FspAI/FastDigest FspAI* (RTGC^GCAC)
  • FspAI/FastDigest FspAI* (RTGC^GCAT)
  • NsbI (FspI)/FastDigest FspI (NsbI) (TGC^GCA)
  • HhaI/FastDigest HhaI
  • Hin6I (HinP1I)/FastDigest HinP1I (Hin6I)
  • MbiI (BsrBI)/FastDigest BsrBI (MbiI)* (GAG^CGG)
  • MspA1I* (CMG^CGG)
  • CviJI
  • HpaII/FastDigest HpaII
  • MspI (HpaII)/FastDigest MspI
  • PdiI (NaeI)/FastDigest NaeI (PdiI) (GCC^GGC)
  • SatI (Fnu4HI)/FastDigest Fnu4HI (SatI)
  • TauI/FastDigest TauI
  • SrfI (GCCC^GGGC)
  • Cac8I