Pvu I

Кат. номер

10650137001

10650129001

Specificity
Pvu I recognizes the sequence CG°AT *CG and generates fragments with 3-cohesive termini.
Recognition sites: CG°AT*CG
CG°AT*CG
Restriction site: CG°AT*CG
CG°AT*CG
Heat inactivation: No inactivation of Pvu I after incubation at 65 °C for 15 minutes.
Quality
Absence of nonspecific endonuclease activities
1µg DNA is incubated for 16hours in 50µl SuRE/Cut Buffer H with an excess of Pvu I. The number of enzyme units which do not change the enzyme-specific pattern is stated in the certificate of analysis.
Absence of exonuclease activity
Approximately 5µg [3H] labeled calf thymus DNA are incubated with 3µl Pvu I for 4hours at +37°C in a total volume of 100µl 50mM Tris-HCl, 10mM MgCl2, 1mM Dithioerythritol, pH approximately 7.5. Under these conditions, no release of radioactivity is detectable, as stated in the certificate of analysis.
DNA Profile
Number of cleavage sites on different DNAs

• : 3
• X174: 0
• Ad2: 7
• M13mp7: 1
• pBR322: 1
• pBR328: 1
• pUC18: 2
• SV40: 0

Unit Definition
One Unit is the enzyme activity that completely cleaves 1 µg DNA in one hour at +37 °C in a total volume of 25 µl (1x) SuRE/Cut Buffer H.
Analysis Note
Activity in PCR buffer: <5%
Relative activity in PCR mix (Taq DNA Polymerase buffer) is less than 5%. The PCR mix contained DNA, primers, 10 mM Tris-HCl (pH 8.3, 20 °C), 50 mM KCl, 1.5 mM MgCl2, 200 µM dNTPs, 2.5 U Taq DNA polymerase. The mix was subjected to 25 amplification cycles.
Compatible ends
Pvu I generates ends that are compatible with fragments generated by Pac I.
Isoschizomers
Pvu I is an isoschizomer to BspC I and Xor II.

Methylation sensitivity
Pvu I cleavage is not inhibited by overlapping dam-methylation at the site indicated (°) on the recognition sequence, but Pvu I fragments of DNA isolated from dam+ strains are not as readily religated as those isolated from dam- strains. Pvu I is inhibited by 5-methylcytosine at the indicated site (°) and by 4-methylcytosine.
SuRE/Cut Buffer System
The buffer in bold is recommended for optimal activity

• A: 50-75%
• B: 75-100%
• H: 100%
• L: 25-50%
• M: 50-75%

Incubation temperature
+37°C
Unit definition
One Unit is the enzyme activity that completely cleaves 1µg DNA in 1 hour at +37°C in a total volume of 25µl SuRE/Cut Buffer H.

Heat inactivation
Pvu I cannot be heat inactivated by incubating it for 15 minutes at +65°C.
PFGE tested
Pvu I has been tested in Pulsed-Field Gel Electrophoresis (on bacterial chromosomes). For cleavage of genomic DNA (E. coli C 600) embedded in agarose for PFGE analysis, we recommend 10U of enzyme/µg DNA and 4 hour incubation time.
Ligation and recutting assay
Pvu I fragments obtained by complete digestion of 1 µg pBR322 DNA are ligated with 1U T4 DNA Ligase in a volume of 10µl by incubation for 16hours at +4°C in 66mM Tris-HCl, 5mM MgCl2, 5mM Dithiothreitol, 1mM ATP, pH 7.5 (at +20°C) resulting in >80% recovery of 1µg pBR322 DNA fragments.
Subsequent re-cutting with Pvu I yields >95% of the typical pattern of pBR322 DNA x Pvu I fragments.
For life science research only. Not for use in diagnostic procedures.

Quality Level	100,
form	solution
packaging	pkg of 100 U (10650137001 [5 U/µl]), pkg of 500 U (10650129001 [5 U/µl])
Торговая марка	Roche
parameter	37 °C optimum reaction temp.
shipped in	dry ice
storage temp.	20°C

RIDADR

NONH for all modes of transport