Klenow Fragment (3′→5′exo-)

Name: Klenow Fragment (3′→5′exo-)
SKU: EMQZ1421

For research use only. Not intended for any clinical use.

Cat.No.

EMQZ1421

Description

Klenow fragment (3' to 5' exo-) is proteolytic product of E. coli DNA Polymerase I which retains polymerization and 5' to 3' exonuclease activity, but has lost 3' to 5' exonuclease activity with D355A and E357A mutation.

Applications

• DNA sequencing by the Sanger dideoxy method• Second strand synthesis of cDNA;• Synthesis of random primers

Concentration

5,000 U/ml

Reaction Conditions

1×Klenow fragment (3' to 5' exo-) Reaction Buffer: [10 mM Tris-HCl (pH 7.9 at 25°C),50 mM NaCl,10 mM MgCl2,1 mM DTT]; add dNTPs (not supplied with enzyme),incubate at 37°C.

Size/Form

200 U; 1,000 U

Source

A recombinant E.coli strain that carries polA gene fragment with D355A and E357A mutation.

Storage

Store at -20°C

Unit Definition

One unit is defined as the amount of enzyme that will incorporate 10 nmol of dNTP into acid-insoluble material in 30 minutes at 37°C.

Publications

Q & A

Customer Reviews

Customer Q&As

How pure is the Klenow Fragment?

A: Klenow Fragment does not contain DNA endonucleases or RNase.

Does Klenow Fragment have exonuclease activity?

A: No, Klenow Fragment, Exo-, lacks exonuclease activity, meaning it retains the 5'→3' polymerase activity of DNA Polymerase I but lacks the intact Klenow enzyme's 5'→3' and 3'→5' exonuclease activities.

Can it generate blunt ends using this enzyme?

A: No, Klenow Fragment, Exo-, lacks exonuclease activity, which can lead to the addition of one or more nucleotides at the 3' end during end-filling, making it unsuitable for generating blunt ends for subsequent ligation.

Where does Klenow Fragment come from?

A: Klenow Fragment is expressed in Escherichia coli, and the expression gene originates from a mutated polA gene fragment.

How to inactivate the Klenow Fragment?

A: Heating at 75°C for 10 minutes or adding an appropriate amount of EDTA can inactivate Klenow Fragment, Exo-.

How to inhibit the activity of the Klenow Fragment?

A: Metal ion chelators, inorganic pyrophosphate (PPi), and high doses of inorganic phosphate (Pi) all have inhibitory effects on Klenow Fragment, Exo-.

How should Klenow Fragment be stored?

A: When using the enzyme, Klenow Fragment should be placed in an icebox or on an ice bath, and after use, it should be immediately stored at -20°C.

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