ADN-ul ce este supus digestiei nu trebuie sa contina contaminati cum ar fi fenol, cloroform, alcool, EDTA, detergenti sau saruri in exces. Se recomanda etape de spalare suplimentare in timpul purificarii. Metilarea ADN-ului poate inhiba digestia cu anumite enzime.
Se pastreaza pe gheata cand nu se afla in congelator. Trebuie adaugata drept ultima componenta in reactie. In general, se recomanda 5-10 unitati de enzima per ug de ADN si 10-20 unitati pentru ADN genomic realizand digestia intr-o ora.
|Unit Definition:||One unit is defined as the amount of enzyme required to digest 1 Ƚg of pBR322 in 1 hour at 37°C in total reaction volume of 50 Ƚl.|
|Shelf Life:||24 months|
|Storage Temp:||-20 °C|
|Storage Conditions:||250 mM NaCl, 10 mM Tris-HCl (pH 7.4), 1 mM DTT, 0.1 mM EDTA, 50% Glycerol, 0.15% Triton X-100, 200 µg/ml BSA|
|Specification Version:||PS-R0616S/L v1.0|
|Exonuclease Activity (Radioactivity Release)||- A 50 µl reaction in CutSmart™ Buffer containing 1 µg of a mixture of single and doublestranded [ ³H] E. coli DNA and a minimum of 30 units of Hpy166II incubated for 4 hours at 37ºC releases|
|Ligation and Recutting (Terminal Integrity)||- After a 10-fold over-digestion of pBR322 DNA with Hpy166II, >95% of the DNA fragments can be ligated with T4 DNA ligase in 16 hours at 16ºC. Of these ligated fragments, >95% can be recut with Hpy166II.|
|Non-Specific DNase Activity (16 Hour)||- A 50 µl reaction in CutSmart™ Buffer containing 1 µg of pBR322 DNA and a minimum of 50 units of Hpy166II incubated for 16 hours at 37ºC results in a DNA pattern free of detectable nuclease degradation as determined by agarose gel electrophoresis.|
|Protein Purity Assay (SDS-PAGE)||- Hpy166II is >95% pure as determined by SDS PAGE analysis using Coomassie Blue detection.|