Technical Data
XapI (ApoI)
Molecular Biology Storage: -20CShipping: Blue Ice
5'-Pu^A A T T Py-3'
3'-Py T T A A^Pu-5'

Xanthomonas ampelina Slo 51021


Supplied as a liquid (Storage Buffer) in 10mM Tris-HCl, pH 7.4, 100mM KCl, 1mM DTT, 1mM EDTA, 0.2mg/ml BSA and 50% glycerol.

Restriction Enzyme Buffer A, 10X (for 100% digestion) R1625:
33mM Tris-acetate, pH 7.9, 10mM magnesium acetate, 66mM potassium acetate and 0.1mg/ml BSA. Incubate at 37C.

Unit Definition:
One unit is defined as the amount of XapI required to digest 1ug of lambda DNA in 1 hour at 37C in 50ul of assay buffer..

Storage and Stability:
May be stored at 4C for short-term only. For long-term storage, store at -20C. Aliquots are stable for at least 6 months at -20C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap. Further dilutions can be made in assay buffer.
Overdigestion Assay:
No detectable change in the specific fragmentation pattern is observed after 15-fold overdigestion with XapI(15u/ug lambda DNA x 1 hours) .

Ligation/Recutting Assay:
After 10-fold overdigestion (5u/ug DNA x 2 hours) with HpaII, more than 95% of the DNA fragments can be ligated at a 5'-termini concentration of 0.2uM. More than 95% of these can be recut.

Labeled Oligonucleotide (LO) Assay: No detectable degradation of single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10 units of XapI for 4 hours.

Blue/White Cloning Assay:
pUC57 was digested at a unique site with 10 units of XapI for 1 hours. After religation and transformation 0.2 % of white colonies were detected.

Star Activity: An excess of XapI (20u/ug DNA x 1 hour) may result in star activity.

Stability during Prolonged Incubation: A minimum of 0.1 units of XapI is required for complete digestion of 1ug of lambda DNA in 16 hours at 37C.

Digestion of Agarose-embedded DNA: A minimum of 5 units of XapI is required for digestion of 1ug of agarose-embedded lambda DNA in 16 hours.

Compatible Ends:
EcoRI, MunI, TasI

Number of Recognition Sites in DNA:
Lambda: 58PhiX174: 5
pBR322: 1
pUC57: 1
pUC18/19: 1
pTZ19R/U: 3
M13mp18/19: 11

Protocol for Digestion:
Add: 44ul nudlease free dH2O
5ul Restriction Enzyme Buffer A, 10X
1ul DNA (0.5-1ug/ul)
0.5-2ul XapI
Mix gently and spin down for a few seconds.
Incubate at 3C for 1-2 hours.

Protocol for Digestion of PCR products Directly after Amplification:
Add: 10ul (~1ug DNA)
16ul nuclease free dH2O
2ul Restriction Enzyme Buffer A, 10X
1-2ul XapI
Mix gently and spin down for a few seconds.
Incubate at 3C for 1-16 hours.

Thermal Inactivation:
XapI is inactivated by incubation at 65C for 20min.
Intended for research use only. Not for use in human, therapeutic, or diagnostic applications.