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FAQs for HaeIII Methyltransferase FAQs for DNA Modifying Enzymes and Cloning Technical Reference for DNA Modifying Enzymes and Cloning Enzyme Finder NEBcutter NEBuffer Chart Double Digest Finder Isoschizomers DNA Sequences and Maps REBASE S-adenosylmethionine (SAM)

Home > Products > DNA Modifying Enzymes and Cloning > Methyltransferases > HaeIII Methyltransferase HaeIII Methyltransferase Catalog # Size Concentration Price Qty   M0224L 2,500 units 10,000 units/ml $244.00 M0224S 500 units 10,000 units/ml $61.00 Prices are in US dollars and valid only for US orders. Download: MSDS PDF Methylation Site: Description: HaeIII Methyltransferase modifies the internal cytosine residue (C5) of the sequence GGCC. Source: A E. coli strain that carries the cloned HaeIII modification gene from Haemophilus aegyptius (ATCC 11116) Reagents Supplied: HaeIII Methyltransferase Reaction Buffer (10X) S-adenosylmethionine (SAM) Reaction & Storage Conditions Reaction Conditions: 1X HaeIII Methyltransferase Reaction Buffer Supplemented with 80 μM S-adenosylmethionine (SAM) Incubate at 37°C. 1X HaeIII Methyltransferase Reaction Buffer: 50 mM Tris-HCl 50 mM NaCl 10 mM Dithiothreitol pH 8.5 @ 25°C Unit Definition: One unit is defined as the amount of enzyme required to protect 1 µg λ DNA in 1 hour at 37°C in a total reaction volume of 10 µl against cleavage by HaeIII restriction endonuclease. Protection Assay Conditions: HaeIII Methyltransferase is incubated with 1 µg of λ DNA in 10 µl of 1X HaeIII Methyltransferase Reaction Buffer, supplemented with 80 µM S-adenosylmethionine, for one hour at 37°C followed by 15 minutes at 65°C. The extent of protection is determined by the addition of 40 µl NEBuffer 2 and 10 units of HaeIII restriction endonuclease. Incubation for 1 hour at 37°C is followed by analysis on an agarose gel. Concentration: 10,000 units/ml Storage Conditions: 50 mM Tris-HCl 50 mM KCl 1 mM Dithiothreitol 10 mM EDTA 200 µg/ml BSA 50% Glycerol pH 7.5 @ 25°C Storage Temperature: -20°C Notes General notes: Storage of SAM: S-adenosylmethionine is stored at –20°C as 32 mM solution dissolved in sulfuric acid (0.005 M) and 10% ethanol. SAM in this solution stored under ideal conditions remains active for up to 6 months. SAM is unstable at (pH 7.5), 37°C, and should be replenished for reactions incubated longer than 4 hours. Many problems in achieving complete digestion can be alleviated by using fresh SAM. HaeIII Methyltransferase protects DNA against cleavage by NotI. FAQs Is S-adenosylmethionine (SAM) supplied with the Methyltransferase? What should be considered if the methylation is not going to completion? Does HaeIII Methyltransferase require DTT? Can HaeIII Methyltransferase be heat inactivated? Does HaeIII Methyltransferase require MgCl2? What is the molecular weight of HaeIII Methyltransferase? Quality Control for Current Lot Quality control values for a specific lot can be found on the datacard which accompanies each product. Endonuclease Activity: Incubation of a 50 μl reaction containing 100 units of HaeIII Methyltransferase with 1 μg of ΦX174 RF I DNA for 4 hours at 37ºC resulted in < 5% conversion to RFII as determined by agarose gel electrophoresis. 16-Hour Incubation:: Incubation of 100 units of HaeIII Methyltransferase with 1 μg of HindIII-digested λ DNA in 50 μl of 1X NEBuffer 2 for 16 hours at 37°C resulted in no detectable endonuclease contamination. Exonuclease Activity:: Incubation of 100 units of HaeIII Methyltransferase with 1 μg sonicated 3H DNA (105 cpm/μg) for 4 hours at 37°C in 50 μl NEBuffer 2 released < 0.05% of the total radioactivity. References Hoffman, J.L. (1986) Biochemistry, 25, 4444-4449. Reagents Sold Separately S-adenosylmethionine (SAM)