SPEI-RO

Spe I

from Sphaerotilus species

• Synonym(e): Spe I, SPE I
• UNSPSC-Code: 12352204

Eigenschaften

• Biologische Quelle: bacterial (Sphaerotilus spp.)
• Qualitätsniveau: 100
• Form: solution
• Spezifische Aktivität: 10000 U/mL
• Verpackung: pkg of 1,000 U (11008951001 [10 U/μl]); pkg of 1,000 U (11207644001 [40 U/μl]); pkg of 200 U (11008943001 [10 U/μl])
• Hersteller/Markenname: Roche
• Parameter: 37 °C optimum reaction temp.
• Farbe: colorless
• pH-Wert: 8.0 (39 °F)
• Löslichkeit: water: miscible
• Eignung: suitable for molecular biology
• Anwendung(en): life science and biopharma; sample preparation
• Fremdaktivität: Endonucleases 10 units, none detected
• Versandbedingung: dry ice
• Lagertemp.: −20°C

Beschreibung

Allgemeine Beschreibung

Spe I recognizes the sequence *A↓*CTAGT and generates fragments with 5′-cohesive termini.

Spezifität

Recognition sites: *A*CTAGT
*A*CTAGT
Restriction site: *A↓*CTAGT
*A↓*CTAGT
Heat inactivation: Spe I can be heat inactivated by incubation at 65 °C for 15 minutes (up to 100 U/μg DNA).

Qualität

Absence of nonspecific endonuclease activities
1μg Ad2 DNA is incubated for 16 hours in 50μl SuRE/Cut Buffer H with an excess of Spe 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 Spe I for 4 hours 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-Profil

Number of cleavage sites on different DNAs

• λ: 0
• φX174: 0
• Ad2: 3
• M13mp7: 0
• pBR322: 0
• pBR328: 0
• pUC18: 0
• SV40: 0

Einheitendefinition

One unit is the enzyme activity that completely cleaves 1 μg Ad2 DNA in one hour at +37 °C in a total volume of 25 μl SuRE/Cut Buffer H.

Lagerung und Haltbarkeit

Do not store below −25°C

Hinweis zur Analyse

Compatible ends
Spe I ends are compatible with ends generated by Bln I, Nhe I, and Xba I.

Isoschizomers
The enzyme is an isoschizomer of Bcu I and Ahl I.

Methylation sensitivity
As indicated by (*) on the recognition sequence above, Spe I is inhibited by the presence of N6-methyladenine and 5′-methylcytosine ( ^mA↓^m CTAGT).

Incubation temperature
+37°C

PFGE tested
Spe 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 using 10U of enzyme/μg DNA and 4 hour incubation time.

Ligation and recutting assay
Spe I fragments obtained by complete digestion of 1μg Ad2 DNA are ligated with 1U T4 DNA Ligase in a volume of 10μl by incubation for 16 hours at +4°C in 66mM Tris-HCl, 5mM MgCl₂, 5mM Dithiothreitol, 1mM ATP, pH 7.5 (at +20°C), resulting in >90% recovery of Ad2 DNA.
Subsequent re-cutting with Spe I yields >95% of the typical pattern of Ad2 × Spe I fragments.

SuRE/Cut Buffer System
The buffer in bold is recommended for optimal activity

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

Activity in PCR buffer: Not tested

Sonstige Hinweise

For life science research only. Not for use in diagnostic procedures.

Nur Kit-Komponenten

• Enzyme Solution
• SuRE/Cut Buffer H 10x concentrated

Sicherheitshinweise

• Lagerklassenschlüssel: 12 - Non Combustible Liquids
• WGK: WGK 1
• Flammpunkt (°F): does not flash
• Flammpunkt (°C): does not flash