biotechrabbit | DNA Blunting Kit

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DNA Blunting Kit

Features

Rapid blunting reaction
Excellent performance in blunt-ended ligation
Ready-to-use blunted product in subsequent ligation

Applications

Blunting and phosphorylation of double-stranded DNA
Preparation of restriction enzyme digested DNA or sheared DNA for blunt-ended ligation

Purchase DNA Blunting Kit

Availability: In stock Bulk requests please inquire at oem@biotechrabbit.com

Catalog #	Size	Package Content	Price*	Qty
BR1101101	100 reactions of 25 µl	125 µl 20X Blunting Enzyme Mix 250 µl 10X Bunting Buffer 25 µl 10 mM dNTP Mix	€348.00

*Does not include applicable tax, shipping & handling, or other charges.

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biotechrabbit™ DNA Blunting Kit shows excellent performance in blunting and phosphorylation of double-stranded DNA. The kit is compatible with a wide variety of dsDNA templates such as restriction enzyme digested DNA fragments, plasmid DNA, PCR products with dA-overhang, sheared or nebulized DNA. The kit is optimized for rapid blunting reaction.

Component	Composition
Blunting Enzyme Mix	20× Blunting Enzyme Mix, in storage buffer containing 50% (v/v) glycerol.
Blunting Buffer	Optimized 10× Blunting Buffer for rapid blunting reaction.
10 mM dNTP Mix	dNTP Mix containing 10 mM dATP, dCTP, dGTP, dTTP in water.

STORAGE

-20°C (until expiry date – see product label)

Quality Control

Functional Assay

Purified double digested sticky-end DNA is blunted, ligated and successful ligation is confirmed.

Exonuclease Activity

Linearized lambda/HindIII DNA fragments are incubated with the enzyme in a 50 µl reaction mixture for 4 h at 37°C. No DNA degradation observed.

Endonuclease/Nick Activity

Supercoiled plasmid DNA is incubated with the enzyme in a 50 µl reaction mixture for 4 h at 37°C. No conversion of covalently closed circular DNA to nicked DNA detected.

Contamination with E. coli DNA

Absence of E. coli genomic DNA is confirmed by qPCR using a sample of the enzyme and specific primers targeting the E. coli 16S rRNA gene. No contamination detected.

Blunting reaction

Scale the reaction volume up as required.
Prepare the blunting reaction as shown below:

Component	Volume	Final concentration
10× Blunting Buffer	2.5 µl	1×
Purified DNA	Variable	up to 5 µg
10 mM dNTP Mix	0.25 µl	0.1 mM
20× Blunting Enzyme Mix	1.25 µl	1×
Nuclease free water	Variable
Total volume	25 µl

Mix and centrifuge briefly to collect the liquid in the bottom of the tube.
Incubate at 25°C (room temperature) for 15 minutes.
For better efficiency or sheared/nebulized DNA extend the incubation time to 30 min.
Heat inactivate the enzyme at 70°C for 10 minutes.
Blunted DNA can be directly used in ligation step (overnight ligation suggested).

Product manual