DpnII (Restriction Enzyme)

REF: EG15533S

Recognition & Cleavage

Recognition site: 5'-GATC-3' (palindromic)

Cleavage pattern:

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

Type: II restriction endonuclease · Common use: Routine cloning, restriction mapping, PCR cleanup

Isoschizomers: MboI, Sau3AI, Bsp143I, BssMI, BstMBI, Kzo9I, NdeII, BstKTI (methylation sensitivities may differ)

Specs

Optimal temperature: 37 °C
Fast digest: 5–15 min in 1× CutOne® / CutOne® Color Buffer
Heat inactivation: 80 °C · 20 min
Storage: −20 °C

Pack Components

Component	EG15533S
DpnII (10 U/µl)	50 µl (500 U)
10× CutOne® Buffer	1 ml
10× CutOne® Color Buffer	1 ml

Recommended Reaction

Buffer: 1× CutOne® or 1× CutOne® Color Buffer
Incubation: 37 °C
Typical duration: 15 min (plasmid); 15–30 min (PCR); 30–60 min (genomic DNA)

CutOne® Color Buffer includes density reagent with red/yellow tracking dyes for direct gel loading.

Quick Protocol (Fast DNA Digestion)

20 µl (plasmid): 15 µl ddH₂O, 2 µl 10× buffer, ≤1 µg DNA, 1 µl DpnII.
30 µl (PCR product): 16 µl ddH₂O, 3 µl 10× buffer, ~0.2 µg DNA, 1 µl DpnII.
If PCR not purified, use 2 µl 10× buffer (residual metal ions).
50 µl (genomic DNA): 30 µl ddH₂O, 5 µl 10× buffer, 10 µl DNA (5 µg), 5 µl DpnII.
Mix gently, quick-spin; incubate at 37 °C for the indicated time.
(Optional) Heat-inactivate at 80 °C for 20 min or load gel directly (Color Buffer).

Multiple Digests

Use ~1 µl of each enzyme; total enzyme volume ≤10% of reaction.
If temperatures differ, start at the lower temperature, then shift higher.
Increase incubation time if total volume >20 µl.

Quality Control

Functional test: 1 µg λDNA (Dam−) + 1 µl DpnII → complete digest in 15 min at 37 °C (20 µl).
Prolonged incubation/star activity: 3 h at 37 °C shows no nuclease degradation; longer incubation may show star activity.
Ligation & recutting: After 10× over-digest, >95% ligation at 22 °C (T4 DNA ligase); >95% of ligated fragments recut.
Non-specific endonuclease: <10% nicked/linearized plasmid after 4 h at 37 °C.
Epigenetics: EpiMark-validated for methylation studies.

Methylation Effects

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

Recognition Sites in Common DNAs

DNA	# Sites	DNA	# Sites
λDNA	116	ΦX174	0
pBR322	22	pUC57	15
pUC18/19	15	SV40	8
M13mp18/19	7	Adeno2	87

Activity in Different Buffers

Buffer	Activity
CutOne®	100%
Thermo FastDigest	75%
NEB rCutSmart™	100%
Takara QuickCut™	75%

*Activities per the functional test conditions above.

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DpnII

Brand: Best Enzymes
Product CAT#: EG15533S
Availability: In Stock

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