McrBC

Application Features

CpG methylation status:
McrBC is a tool for determining the methylation state of CpG dinucleotides (6-10). McrBC will act upon a pair of Pu^mCG sequence elements, thereby detecting a high proportion of methylated CpGs, but will not recognize HpaII/MspI sites (CCGG) in which the internal cytosine is methylated.
Detection of cytosine-methylated DNA:
The very short half-site consensus sequence (Pu^mC) allows a large proportion of the methylcytosines present to be detected. Even DNA which is not heavily methylated can be detected, as a low level of cleavage occurs even when the Pu^mC elements are as far as 3 kb apart.
Enrichment for undermethylated DNA (11).

Companion Products

• Monarch® Plasmid Miniprep Kit 
• Monarch® DNA Gel Extraction Kit
• Monarch® PCR & DNA Cleanup Kit (5 μg)

Materials Sold Separately

• NEBuffer 2
• Recombinant Albumin, Molecular Biology Grade
  
  
  
  
  

1. McrBC makes one cut between each pair of half-sites, cutting close to one half-site or the other, but cleavage positions are distributed over several base pairs approximately 30 base pairs from the methylated base (2). Therefore, the enzyme does not produce defined DNA ends upon cleavage. Also, when multiple McrBC half-sites are present in DNA (as is the case with cytosine-methylated genomic DNA) the flexible nature of the recognition sequence results in an overlap of sites, and so a smeared rather than a sharp banding pattern is produced.
2. McrBC cleavage of the supplied 4.3 kb linear, methylated control plasmid DNA produces several fragments between approximately 700 bp and 2.3 kb in size.
3. GTP is more labile than other nucleotides. We recommend aliquoting the 100 mM solution supplied and thawing and diluting as necessary.

1. Sutherland, E. et al. (1992). J. Mol. Biol.. 225, 327-334.
2. Chotai, K.A. and Payne, S.J. (1998). J. Med. Genet.. 35, 472-475.
3. Burman, R.W. et al. (1999). Am. J. Hum. Genet.. 65, 1375-1386.
4. Santoso, B. et al. (2000). J. Biol. Chem.. 275, 1952-1958.
5. Lyko, F. et al. (2000). Nat. Genet.. 23, 363-366.
6. Gowher, H. et al. (2000). EMBO J.. 19, 6918-6923.
7. Zhou, Y. et al. (2002). Genome. 45, 91-99.
8. Irizarry, R.A. et al. (2008). Genome Res.. 18, 780-790.
9. Hublarova, P. et al. (2009). Int J Gynecol Cancer. 19, 321-325.
10. Stewart, F.J. and Raleigh, E.A. (2009). Biol. Chem.. 379, 611-616.
11. Panne, D. et al. (1999). J. Mol. Biol.. 290, 49-60.
12. Stewart, F.J. et al. (1999). J. Mol. Biol.. 298, 611-622.
13. Raleigh, E.A. (1992). Mol. Microbiol.. 6, 1079-1086.

1. Protocol for cleavage with McrBC (M0272)

• Activity of DNA Modifying Enzymes in rCutSmart™ Buffer

• DNA Methylation Table
• Restriction Enzymes for Epigenetics Selection Chart

1. Does McrBC cut hemi-methylated DNA?
2. Does McrBC produce blunt or sticky ends?
3.  Why does my McrBC cleaved DNA smear when run on an agarose gel?
4. How much enzyme should be used for cleaving genomic DNA?
5. Is extended digestion of McrBC recommended?
6. Are there any published papers in which McrBC has been used?
7. Will McrBC work in rCutSmart Buffer?

Quality Control tests are performed on each new lot of NEB product to meet the specifications designated for it. Specifications and individual lot data from the tests that are performed for this particular product can be found and downloaded on the Product Specification Sheet, Certificate of Analysis, data card or product manual. Further information regarding NEB product quality can be found here.

The Specification sheet is a document that includes the storage temperature, shelf life and the specifications designated for the product. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]

• M0272S_L_v1

The Certificate of Analysis (COA) is a signed document that includes the storage temperature, expiration date and quality controls for an individual lot. The following file naming structure is used to name these document files: [Product Number]_[Size]_[Version]_[Lot Number]

• M0272S_L_v1_0351305
• M0272S_L_v1_0351306
• M0272S_L_v1_0351307
• M0272S_L_v1_0351409
• M0272S_L_v1_0351412
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• M0272S_v1_10185392
• M0272L_v1_10200381
• M0272S_v1_10197160

The following is a list of Safety Data Sheet (SDS) that apply to this product to help you use it safely.

• McrBC

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