Restriction endonucleases

Description

Product Details

Asu II is a restriction enzyme purified from an isolated strain (#94S).

Reagents supplied: 10x AsuII and 10x EQ buffer

Unit definition: One unit is the amount of enzyme required to digest 1 μg of Lambda DNA (Hind III digest) in 60 min at 37^oC in a total reaction volume of 50 μl.

Digest Guidelines

• Standard reaction conditions: In order to overcome DNA source variability, a ~10 fold overdigest is suggested:

10U Asu II	1μl
10x EQ buffer	5μl
DNA substrate	1μg
Sterile ultrapure water	Up to 50 μl

• Speed Digest reaction: Incubate for 15 minutes at 37^oC as in standard reaction in 1x AsuII buffer (50 mM NaCl, 10 mM Tris-HCl (pH 7.9 @ 25°C), 10 mM MgCl2, 0.1% Triton X-100, 100 μg/ml BSA ) or  1x EQ buffer (20 mM Tris-acetate (pH 7.9 @ 25^oC), 10 mM Mg-acetate, 50 mM K-acetate, 100 μg/ml BSA).

• Activity(%) in EnzyQuest buffers -Double digest tips

R1	R2	R3	R4	EQ
75	100	50-75	25	100

Reactions were performed as for unit definition reaction. The above data could be used as guideline for double digest reactions. Our enzymes are also active and compatible in buffer systems from other vendors.

Heat inactivation: 65^oC for 20 minutes

Methylation Sensitivity:

• dam methylation: Not sensitive
• dcm methylation: Not sensitive
• CpG methylation: Blocked

Unit calculation assay conditions:

• 1x AsuII buffer
• Lambda DNA (Hind III digest)
• Incubation at 37^oC

Storage conditions:

• 100 mM KCl
• 10 mM Tris-HCl (pH 7.9 @25^oC)
• 0.1 mM EDTA
• 1 mM dithiothreitol
• 0.15% Triton X-100
• 200 μg/ml BSA
• 50% glycerol
• Store at -20^oC

Non-specific nuclease activity: 100 units of Asu II do not produce any unspecific cleavage products after 16 hrs incubation with 1 μg of Lambda DNA (Hind III digest) at 37^oC.

Re-ligation and recut: After 50-fold overdigestion with Asu II, greater than 95% of the DNA fragments can be ligated and recut with this enzyme.