Unit Definition:One unit is defined as the amount of this enzyme required to digestcompletely 1ug of N6-methyladenine free lambda DNA in 50ul of the reaction mixture at 37ºC for 1 hour.
Form:Supplied as a liquid in 10mM Tris-HCl, pH 7.5, 100mM KCl, 0.1mM EDTA, 1mM DTT, 0.15% Triton X-100, 0.01mg/ml BSA, 50% glycerol
Supplied with:551600A: Loading Buffer, 10X:Dilute to 1X for use. 10X buffer composition is 0.9% SDS, 0.05% Bromophenol Blue, 50% glycerol. Add Loading Buffer, 1X to stop enzyme reaction and apply on agarose gel electrophoresis. SDS may precipitate during the storage at RT. The loading buffer, 1X can be stored at RT once used. If precipitate forms, dissolve in a warm water batch before use.
Supplied with 551600B L Buffer, 10X:Dilute to 1X for use. 10X buffer composition is 100mM Tris-HCl pH 7.5, 100mM MgCl2, 10mM Dithiothreitol
Effect of DNA Methylation:Enzyme activity is affected by dam methylase, therefore general DNA originated from E. coli cannot be cleaved by this enzyme.
Reaction Temperature:37°C
Quality Control Data:Overdigestion Assay:Non-specific nuclease activity was not detected after incubating 1ug of N6-methyladenine free lambda DNA with 15 units of this enzyme in 50ul of the buffer overnight at 37ºC, as assessed by agarose gel electrophoresis.
Ligation and Recleavage (L/R) Assay:After 10-fold overdigestion with this enzyme, the DNA fragments caould be ligated with T4 DNA lipase and recut with this enzyme. Ligation: >95%, Recutting: >95%
Labeled Oligonucleotide (LOA) Assay:The degradation rate of oligonucleotides was less than 10% during incubation with 6 U of this enzyme for 1 hour in 10ul of the buffer.
Number of Recognition Sites in DNA:Lambda: 21Ad2: 22SV40: 3PhiX174: 0pBR322: 8pUC19: 7pUC119: 7M13mp18/19: 3Col E1: 4
General Reaction Mixture:MflI 1ul551600B2ulSubstrate DNA≤1ugNuclease-free water up to 20ul
Storage and Stability:Aliquot to avoid repeated freezing and thawing and store at -20°C. Aliquots are stable for 12 months after receipt. For maximum recovery of product, centrifuge the original vial after thawing and prior to removing the cap.
