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 realizandu-se digestia intr-o ora.
Product Name | HpyCH4III |
Concentration: | 5,000 units/ml |
Unit Definition: | One unit is defined as the amount of enzyme required to digest 1 µg of Lambda DNA in 1 hour at 37°C in a total reaction volume of 50 µl. |
Shelf Life: | 24 months |
Storage Temp: | -20 °C |
Storage Conditions: | 100 mM NaCl, 10 mM Tris-HCl (pH 7.4), 1 mM DTT, 0.1 mM EDTA, 50% Glycerol, 200 µg/ml BSA |
Specification Version: | PS-R0618S/L v1.0 |
Effective Date: | 23-Sep-13 |
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 15 units of HpyCH4III incubated for 4 hours at 37ºC releases |
Ligation and Recutting (Terminal Integrity) | - After a 5-fold over-digestion of Lambda DNA with HpyCH4III, ~50% 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 HpyCH4III. |
Non-Specific DNase Activity (16 Hour) | ) - A 50 µl reaction in CutSmart™ Buffer containing 1 µg of Lambda DNA and a minimum of 5 units of HpyCH4III incubated for 16 hours at 37ºC results in a DNA pattern free of detectable nuclease degradation as determined by agarose gel electrophoresis. |
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