Restriction endonucleases

Description

Product Details

Taq I is a restriction enzyme purified from Thermus aquaticus YT I.

Reagents supplied: 10x TaqI

Unit definition: One unit is the amount of enzyme required to digest 1 μg of Lambda DNA (dam^–) in 60 min at 65^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 Taq I	1μl
10x eQ buffer	5μl
DNA substrate	1μg
Sterile ultrapure water	Up to 50 μl
Incubate for 60 min at 65ºC

• Speed Digest reaction: Incubate for 15 minutes at 65^oC as in standard reaction in 1x TaqI buffer (100 mM KCl, 20 mM Tris-HCl (pH 8.5 @ 25°C), 3 mM MgCl2, 0.04% 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
10-25	50-75	75-100	50-75	–

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: 80ºC for 20 minutes 

Methylation Sensitivity: 

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

 Unit calculation assay conditions:  

• 1x TaqI buffer 
• Lambda DNA (dam-) 
• Incubation at 65ºC  

 Storage conditions:  

• 300 mM KCl  
• 10 mM Tris-HCl (pH 7.4) 
• 0.1 mM EDTA  
• 1 mM dithiothreitol 
• 500 μg/ml BSA 
• 50% glycerol  
• Store at -20ºC 

Non-specific nuclease activity: 100 units of Taq I do not produce any unspecific cleavage products after 16 hrs incubation with 1 μg of Lambda DNA (dam-) at 65ºC. 

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