Source: E.coli strain that carries the cloned Zra I gene from Zoogloea ramigera 11
10 mM Tris-HCl (pH 7.4); 50 mM NaCl; 0,1 mM EDTA; 200 µg/ml BSA; 1 mM DTT; 50% glycerol.
Incubate at 37°C.
10 mM Tris-HCl (pH 7.6 @ 25ºC), 10 mM MgCl2, 1 mM DTT
One unit is defined as the amount of enzyme required to digest 1 µg of λ DNA in 1 hour at 37°C in a total reaction volume of 50 µl.
Quality Control Assays
After 10-fold overdigestion with Zra I, approximately 90% of λ DNA fragments can be ligated with T4 DNA Ligase and recut. In the
presence of 10% PEG ligation is better.
A 50 µl reaction containing 1 µg of λ DNA and 10 units of enzyme incubated for 16 hours resulted in the same pattern of DNA bands as a reaction
incubated for 1 hour.
No detectable degradation of a single- and double-stranded oligonucleotide was observed after incubation with 10 units of enzyme for 3 hours.
Activity in SEBuffers:
SEBuffer B 100%
SEBuffer G 50-75%
SEBuffer O 25-50%
SEBuffer W 25-50%
SEBuffer Y 75-100%
SEBuffer ROSE 100%
When using a buffer other than the optimal (suppied) SEBuffer, it may be necessary to add more enzyme to achieve complete digestion.
Zra I is a neoschizomer of Aat II.
Yes (80°C for 20 minutes)
Reagents supplied with enzyme:
High enzyme concentration may result in star activity. The minimum number of units that resulted in complete digestion of 1 ug of substrate DNA in
16 hours is 0,5. Zra I cleaves linear plasmid DNA at a rate 1,5-2 times higher than supercoiled plasmid DNA.