NdeI

Molecular Biology / MB-Enzymes, Restriction

-20°C

Supplied as a liquid in 10mM Tris-HCl (pH 7.4 at 25°C), 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA and 50% glycerol.

5'-C A^T A T G-3'
3'-G T A T^A C-5'

Source:
Neisseria denitrificans

Concentration:
~10u/ul

Buffers:
R1625-Restriction Enzyme Buffer A (double digests): 1X Buffer composition-Supplied as a liquid containing 33mM Tris-acetate pH 7.9, 10mM magnesium acetate, 66mM potassium acetate, 0.1mg/ml BSA. Incubate at 37°C.

R1625-03-Restriction Enzyme Buffer D (for 100% digestion):
1X Buffer composition-Supplied as a liquid in 50mM Tris-HCl, pH 7.5, 10mM MgCl2, 100mM sodium chloride, 0.1mg/ml BSA. Incubate at 37°C.

Overdigestion Assay:
No detectable change in the specific fragmentation pattern is observed after 160-fold overdigestion (10u/ug lambda DNA x 16 hours) with NdeI.

Ligation/Recutting Assay:
After 50-fold overdigestion (3u/ug DNA x 17 hours) with NdeI, more than 95% of the DNA fragments can be ligated at a 5'-termini concentration of 0.1uM. More than 95% of these can be recut.

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

Stability during Prolonged Incubation:
A minimum of 0.3 units of Ndel is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.

Thermal Inactivation:
Ndel is inactivated by incubation at 65°C for 20min.

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

Compatible Ends:
Csp6I, FspBI, Tru1I, VspI

Number of Recognition Sites in DNA:
Lambda: 7
PhiX174: 0
M13mp18/19: 3
pBR322: 1
pUC18/19: 1
pUC57: 1
pTZ19R/U: 0

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

Applications:
• Molecular cloning
• Restriction site mapping
• Genotyping
• Southern blotting
• Restriction fragment length polymorphism (RFLP)
• SNP

Dilution Buffer:
10mM Tris-HCl, pH 7.4 at RT, 100mM KCl, 1mM EDTA, 1mM DTT, 0.2mg/ml BSA and 50% glycerol.

Enzyme Activity in Buffer A (R1625)/Buffer D (R1625-03):
R1625-0-20%
R1625 (2X)-50-100%
R1625-03-100%

Storage and Stability:
May be stored at 4°C for short-term only. Aliquot to avoid repeated freezing and thawing. Store at -20°C. Aliquots are stable for 6 months after receipt at -20°C. 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.