SfiI (Restriction Enzyme)

REF: EG15570S

Recognition & Cleavage

Recognition site: 5'-GGCCNNNNNGGCC-3'

Cleavage pattern:

5' ... G G C C N N N N^▼N G G C C ... 3'
3' ... C C G G N_▲N N N N C C G G ... 5'

Type: II restriction endonuclease · Common use: Rare cutter for cloning, mapping, and DNA assembly

Specs

Optimal temperature: 50 °C
Fast digest: 5–15 min in 1× CutOne® / CutOne® Color Buffer
Heat inactivation: Not heat-inactivatable (purify DNA after digestion)
Storage: −20 °C

Pack Components

Component	EG15570S
SfiI (20 U/µl)	100 µl (2000 U)
10× CutOne® Buffer	1 ml
10× CutOne® Color Buffer	1 ml

Recommended Reaction

Buffer: 1× CutOne® or 1× CutOne® Color Buffer
Incubation: 50 °C
Typical duration: 15 min (plasmid DNA); 15–30 min (PCR); 30–60 min (genomic DNA)

SfiI cannot be heat-inactivated. After digestion, purify DNA with spin column or phenol/chloroform before downstream use.

Quick Protocol (DNA Digestion)

20 µl (plasmid): 15 µl ddH₂O, 2 µl 10× CutOne®, ≤1 µg DNA, 1 µl SfiI.
30 µl (PCR product): 16 µl ddH₂O, 3 µl 10× CutOne®, ~0.2 µg DNA, 1 µl SfiI.
If PCR not purified, reduce buffer to 2 µl.
50 µl (genomic): 30 µl ddH₂O, 5 µl 10× CutOne®, 5 µg DNA, 5 µl SfiI.
Mix gently, spin down; incubate at 50 °C for recommended time.
Purify DNA (column or phenol/chloroform). If Color Buffer used, load directly to gel for check.

Notes

Keep total enzyme volume ≤10% of reaction (avoid excess glycerol).
Contaminants (phenol, ethanol, EDTA, high salt, detergents) inhibit activity—purify DNA if needed.
Scale incubation time up if reaction volume exceeds 20 µl.

Quality Control

Functional test: Complete digestion of 1 µg pEPE DNA in 15 min at 50 °C.
Prolonged incubation: 3 h at 50 °C shows no nuclease degradation (longer incubation may cause star activity).
Ligation & recutting: >95% ligation efficiency (T4 ligase, 22 °C) and >95% recutting of ligated DNA.
Non-specific endonuclease: <10% nicked/linearized plasmid after 4 h at 50 °C.

Methylation Effects

Methylase	Effect
Dam	No effect
Dcm	Impaired
CpG	Impaired
EcoKI	No effect
EcoBI	No effect

Recognition Sites in Common DNAs

DNA	DNA	# Sites
λDNA	ΦX174	0
pBR322	pUC57	0
pUC18/19	SV40	1
M13mp18/19	Adeno2	3

Buffer Compatibility (Activity)

Buffer	Activity
CutOne®	100%
Thermo FastDigest	100%
NEB rCutSmart™	100%
Takara QuickCut™	100%

*Activities based on functional test conditions.

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SfiI

Brand: Best Enzymes
Product CAT#: EG15570S
Availability: In Stock

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