Overdigestion Assay: No detectable change in the specific fragmentation pattern is observed after 80-fold overdigestion with Mbil (5u/ug lambda DNA x 16 hours).
Ligation/Recleavage Assay: The ligation and recleavage assay was replaced with L0 test after validating experiments showed L0 test ability to trace nuclease and phosphatase activities with sensitivity that is higher than L/R by a factor of 100.
Labeled Oligonucleotide (LO) Assay: No detectable degradation of a single-stranded and double-stranded labeled oligonucleotide was observed after incubation with 10 units of Mbil for 4 hours.
Methylation Effects:MbiI never overlaps Dam, Dcm, EcoKlCpG cleavage impairedEcoBl may overlap
Recommended Protocol for Digestion:Add:Nuclease-free water16ulR16252ulDNA (0.5-1ug/ul)1ulM24570.5-2ul*
Mix gently and spin down for a few seconds.
Incubate at 37ºC for 1-16 hours*The digestion raction may be scaled either up or down
Stability during Prolonged Incubation: A minimum of 0.2 units of MbiI is required for complete digestion of 1ug of lambda DNA in 16 hours at 37°C.
Thermal Inactivation: MbiI is inactivated by incubation at 65°C for 20 minutes.
Digestion of Agarose-embedded DNA: A minimum of 5 units of MbiI is required for digestion of 1ug of agarose-embedded lambda DNA in 16 hours.
Number of Recognition Sites in DNA:Lambda: 17 pUC57: 3PhiX174: 1 pTZ19R/U: 4M13mp18/19: 4 pUC18/19: 3pBR322: 2
Storage and Stability:May be stored at 4°C for short-term only. For long-term storage, 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.
