Restriction endonucleases

Description

Product Details

Digest Guidelines 

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

10U SstI	1μl
10x EQ buffer	5μl
DNA substrate	1μg
Sterile ultrapure water	Up to 50 μl
Incubate for 60 min at 37oC

• Speed Digest reaction:  Incubate for 15 minutes as in standard reaction in 1x R1 or 1x EQ buffer. 

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

R1	R2	R3	R4	R5	EQ
100	25-50	25	<10	50	–

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: 65oC for 20 minutes 

Methylation Sensitivity: 

dam methylation: Not sensitive 

dcm methylation: Not sensitive 

CpG methylation: Not sensitive 

Unit calculation assay conditions:  

• 1x R1 buffer 
• Lambda DNA (Hind III digest) 
• Incubation at 37ºC  

 Storage conditions:  

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

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

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