KLEN-RO

Klenow Enzyme

Labeling grade, from Escherichia coli lysogenic NM 964

• Synonym(s): Klenow
• Enzyme Commission number: 2.7.7.7 (BRENDA, IUBMB)

Properties

• biological source: Escherichia coli
• Quality Level: 100
• form: solution
• specific activity: >5000 units/mg protein
• packaging: pkg of 100 U (11008404001); pkg of 500 U (11008412001)
• manufacturer/tradename: Roche
• storage temp.: −20°C

Description

General description

Klenow enzyme is a DNA-dependent 5′→3′ polymerase with 3′→5′ exonuclease activity. It lacks the 5′→3′ exonuclease activity of the native enzyme. The addition of mononucleotides from dNTPs to the 3′-OH terminus of DNA is catalyzed. This property is used to synthesize DNA complementary to single-stranded DNA templates.

Specificity

Heat inactivation: Add 2 μl 0.2 mM EDTA (pH 8.0) and/or heat to 65 °C for 10 minutes

Packaging

100, 500 units

Application

Use Klenow Enzyme for:

• Random-primed DNA labeling using random oligonucleotides as primers for the incorporation of nonradioactively labeled and ³²P-labeled nucleotides.
• Fill-in reaction for blunt-end formation of 3′-recessed (staggered) ends.

Quality

Absence of endo- and exonuclease activity tested according to the current Quality Control procedures

Unit Definition

One unit is the enzyme activity which incorporates 10 nmol of total nucleotides into an acid-precipitable fraction in 30 minutes under assay conditions at +37 °C with poly [d(A-T)] as primer (Richardson, C.C. & Kornberg, A. (1994) J. Biol. Chem. 244, 2996).

Volume Activity: 2,000 U/ml

Physical form

Enzyme in Storage buffer: 50mM potassium phosphate, 1mM dithioerythritol, 50% glycerol (v/v), pH 7.0 (+4°C).

Other Notes

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

Safety Information

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