PvuII (Restriction Enzyme)

REF: EG15604S

Recognition & Cleavage

Recognition site: 5′-CAGCTG-3′ (palindromic; blunt ends)

Sequence context:

5′ ... C A G^▼C T G ... 3′
3′ ... G T C_▲G A C ... 5′

Type: II restriction endonuclease · Common use: Routine cloning & restriction mapping. :contentReference[oaicite:1]{index=1}

Specs

Optimal temperature: 37 °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	EG15604S
PvuII (20 U/µl)	200 µl (4000 U)
10× CutOne® Buffer	2 × 1 ml
10× CutOne® Color Buffer	2 × 1 ml

Specs & components; heat-inactivation note. :contentReference[oaicite:2]{index=2}

Recommended Reaction

Buffer: 1× CutOne® or 1× CutOne® Color Buffer
Incubation: 37 °C
Guideline: use fast-digestion setup below; scale volumes as needed

Color Buffer includes density reagent + red/yellow dyes for direct gel loading. :contentReference[oaicite:3]{index=3}

Quick Protocol (Fast DNA Digestion)

20 µl (plasmid): 15 µl ddH₂O, 2 µl 10× buffer, ≤1 µg DNA, 1 µl PvuII.
30 µl (PCR product): 16 µl ddH₂O, 3 µl 10× buffer, ~0.2 µg DNA, 1 µl PvuII.
If PCR is unpurified, use 2 µl 10× buffer due to residual metal ions.
50 µl (genomic DNA): 30 µl ddH₂O, 5 µl 10× buffer, 10 µl DNA (5 µg), 5 µl PvuII.
Mix gently, quick-spin; incubate at 37 °C for 15 min (plasmid), 15–30 min (PCR), or 30–60 min (genomic).
Note: This enzyme cannot be heat-inactivated; purify DNA after digestion (column or phenol/chloroform).
If Color Buffer was used, an aliquot can be loaded directly on gel. :contentReference[oaicite:4]{index=4}

Double / Multiple Digests

Use ~1 µl of each enzyme; keep total enzyme volume ≤10% of reaction.
If enzymes require different temperatures, start at the lower temperature then shift higher.
Increase incubation if total volume >20 µl. :contentReference[oaicite:5]{index=5}

Quality Control

Functional test: 1 µg λDNA + 1 µl PvuII in 20 µl CutOne® → complete digest in 15 min at 37 °C.
Prolonged incubation / star activity: 3 h at 37 °C shows no nuclease degradation; longer incubation may cause star activity.
Ligation & recutting: After 10× over-digest, >95% ligation at 22 °C; >95% of ligated fragments recut.
Non-specific endonuclease: <10% nicked/linearized plasmid after 4 h at 37 °C. :contentReference[oaicite:6]{index=6}

Methylation Effects

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

Methylation data. :contentReference[oaicite:7]{index=7}

Recognition Sites in Common DNAs

DNA	# Sites	DNA	# Sites
λDNA	15	ΦX174	0
pBR322	1	pUC57	2
pUC18/19	2	SV40	3
M13mp18/19	3	Adeno2	24

Site counts. :contentReference[oaicite:8]{index=8}

Activity in Different Buffers

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

Activities from the functional test conditions. :contentReference[oaicite:9]{index=9}

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PvuII

Brand: Best Enzymes
Product CAT#: EG15604S
Availability: In Stock

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