ScaI is a restriction enzyme purified from Streptomyces caespitosus.
Reagents supplied: 10x ScaI buffer and 10x eQ buffer
Unit definition: One unit is the amount of enzyme required to digest 1 μg of Lambda DNA in 60 min at 37oC in a total reaction volume of 50 μl.
Standard reaction conditions: In order to overcome DNA source variability, a ~10 fold overdigest is suggested:
10U Sca I
10x ScaI buffer
Sterile ultrapure water
Up to 50 μl
Speed Digest reaction: Incubate for 15 minutes at 37oC as in standard reaction in 1x ScaI buffer (100 mM KCl, 10 mM Bis-Tris-Propane (pH 7.0 @ 25°C), 10 mM MgCl2, 100 μg/ml BSA) or 5 units in 2x eQ buffer (40 mM Tris-acetate (pH 7.9 @ 25oC), 20 mM Mg-acetate, 100 mM K-acetate, 200 μg/ml BSA) .
Activity(%) in EnzyQuest buffers-Double digest tips
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
dam methylation: Not sensitive
dcm methylation: Not sensitive
CpG methylation: Not sensitive
Unit calculation assay conditions:
1x ScaI buffer
Incubation at 37oC
50 mM KCl,
10 mM Tris-HCl (pH 7.4)
0.1 mM EDTA
1 mM dithiothreitol
500 μg/ml BSA
Store at -20oC
Non-specific nuclease activity: 600 units of Sca I do not produce any unspecific cleavage products after 16 hrs incubation with 1 μg of Lambda DNA at 37oC.
Re-ligation and recut: After 100-fold overdigestion with Sca I, greater than 90% of the DNA fragments can be ligated and recut with this enzyme.
Star activity: Large excess of the enzyme may results in the appearance of star activity.
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