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A1372-05 Alw21I (HgiAI) CAS:

Specifications
Grade
Molecular Biology Grade
EU Commodity Code
38220090
Shipping Temp
Blue Ice
Storage Temp
-20°C

5' - G W G C W^C - 3' 3' - C^W C G W G - 5’

Source
Acinetobacter lwoffi RFL21
Concentration
10u/ul
Form
Supplied as a liquid in 10mM Tris-HCl pH 7.5, 300mM sodium chloride, 1mM DTT, 0.1mM EDTA, 0.2mg/ml BSA, 50% glycerol.
Dilution Buffer
10mM Tris-HCl pH 7.4, 100mM potassium chloride, 1mM DTT, 1mM EDTA, 0.2mg/ml BSA, 50% glycerol.
Unit Definition
One unit is defined as the amount of Alw21I required to digest 1ug of lambda DNA in 1 hour at 37°C in 50ul of assay buffer.
Supplied With
R1625: Restriction Enzyme Buffer A, 10X: Supplied as a liquid in 33mM Tris-acetate, 10mM magnesium acetate, 66mM potassium acetate, 0.1mg/ml BSA pH 7.9 at 37°C when diluted to 1X. Note: 1X conc. yields 20-50% enyzme activity. 2X conc. yields 50-100% enzyme activity.
R1625-03: Restriction Enzyme Buffer D, 10X: Supplied as a liquid in 50mM Tris-HCl, pH 7.5, 10mM MgCl2, 100mM sodium chloride, 0.1mg/ml BSA when diluted to 1X. Incubate at 37°C. Note: This buffer yields 100% enzyme activity
Overdigestion Assay
No detectable change in the specific fragmentation pattern is observed after 160-fold overdigestion (10u/ug lambda DNA x 16 hours) with Alw21I.
Ligation/Recutting Assay
After 50-fold overdigestion (3u/ug DNA x 17 hours) with Alw21I, more than 95% of the DNA fragments can be ligated at a 5'-termini concentration of 0.27uM. More than 95% of these sites 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 Alw21I for 4 hours.
Stability During Prolonged Incubation
A minimum of 0.1 units of Alw21I is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.
Compatible Ends
GTGCA^C - BseSI, SduI, Mph1103I, PstI, SdaI GAGCT^C - Eco24I, SacI, SduI GAGCA^C - SduI
Number of Recognition Sites in DNA
Lambda: 28 PhiX174: 3 M13mp18/19: 3 pBR322: 8 pUC18/19: 5 pUC57: 5 pTZ19R/U: 4
Thermal Inactivation
Alw21I is inactivated by incubation at 65°C for 20 min.
Add
Nuclease free water: 16ul 10X Restriction Enzyme Buffer D: 2ul DNA (0.5-1ug/ul): 1ul Alw21I: 0.5-2ul Mix gently, spin down for a few seconds. Incubate at 37ºC for 1-16 hours. This digestion reaction may be scaled either up or down.
Add
PCR Reaction Mixture: 10ul (~0.1-0.5ug of DNA) Nuclease free water: 18ul 10X Restriction Enzyme Buffer D: 2ul Alw21I: 1-2ul Mix gently, spin down for a few seconds. Incubate at 37ºC for 1-16 hours.
Methylation Effects on Digestion
Dam, Dcm: Never overlaps, no effect CpG, EcoKI: May overlap, no effect EcoBI: May overlap, effect not determined
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 at -20°C. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
Source
Acinetobacter lwoffi RFL21
Concentration
~10u/ul
Important Note
This product as supplied is intended for research use only, not for use in human, therapeutic or diagnostic applications without the expressed written authorization of United States Biological.
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