BLNI-RO

Bln I (Avr II)

from Brevibacterium linens

• UNSPSC Code: 12352204

Properties

• biological source: bacterial (Brevibacterium linens)
• Quality Level: 100
• form: solution
• specific activity: 10000 U/mL
• packaging: pkg of 1,000 U (11558170001 [10 U/μl]); pkg of 200 U (11558161001 [10 U/μl])
• manufacturer/tradename: Roche
• parameter: 37 °C optimum reaction temp.
• color: colorless
• pH: 8.1 (39 °F)
• solubility: water: miscible
• suitability: suitable for molecular biology
• application(s): life science and biopharma; sample preparation
• foreign activity: Endonucleases, none detected (up to 20 U with MWM II-DNA); Endonucleases, none detected (up to 20U with pBR 322-DNA)
• shipped in: dry ice
• storage temp.: −20°C

Description

General description

Bln I recognizes the sequence C↓CTAGG and generates fragments with 5′-cohesive termini.

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

Isoschizomers
Bln I is an isoschizomer of Avr II.
Note: The complete 13 site Avr II restriction map of the E.coli genome has been reported.

Methylation sensitivity
The enzyme is not known to be affected by methylation.

Specificity

Recognition sites: CCTAGG
CCTAGG
Restriction site: C↓CTAGG
C↓CTAGG
Heat inactivation: No inactivation of Bln I after incubation at 65 °C for 15 minutes.

Quality

Absence of nonspecific endonuclease activities
1 μg λDNA is incubated for 16 hours in 50 μl SuRE/Cut Buffer H with an excess of Bln 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 5 μg [³H] labeled calf thymus DNA are incubated with 3 μl Bln I for 4 hours at +37°C in a total volume of 100 μl 50 mM Tris-HCl, 10 mM MgCl₂, 1 mM Dithioerythritol, pH approximately 7.5. Under these conditions, no release of radioactivity is detectable, as stated in the certificate of analysis.

Typical ligation and recutting assay
Bln I fragments obtained by complete digestion of 1 μg λ × EcoR I DNA ligated for 16 hours at +4°C with 1 U T4 DNA Ligase in 10 μl buffer that contains 66 mM Tris-HCl, 5 mM MgCl₂, 5 mM Dithiothreitol, 1 mM ATP, pH 7.5 (at +20°C). The percentages of product that can be ligated and subsequently recut with Bln I and EcoR I (yielding the typical pattern of λ × EcoR I × Bln I fragments) are stated under "Lig" and "Rec" in the certificate of analysis.

DNA Profile

Number of cleavage sites on different DNAs

• λ: 2
• φX174: 0
• Ad2: 2
• M13mp7: 0
• M13mp18:0
• pBR322: 0
• pBR328: 0
• pUC18: 0
• SV40: 2

Unit Definition

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

Storage and Stability

Do not store below −25°C.

Analysis Note

PFGE tested
Bln 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 10 U of enzyme/μg DNA and 4 hour incubation.

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

• A: 25-50%
• B: 50-75%
• H: 100%
• L: 0-10%
• M: 25-50%

Activity in PCR buffer: Not tested

Other Notes

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

Kit Components Only

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

Safety Information

• Storage Class Code: 12 - Non Combustible Liquids
• WGK: WGK 1
• Flash Point(F): does not flash
• Flash Point(C): does not flash