Storage: −20°C
UNSPSC Code: 12352200
Components: Enzyme Solution; SuRE/Cut Buffer A 10x concentrated
RIDADR: NONH for all modes of transport
Analysis Note:
Activity in PCR buffer: 100%
Relative activity in PCR mix (Taq DNA Polymerase buffer) is 100%. The PCR mix contained λDNA, primers, 10 mM Tris-HCl (pH 8.3, 20 °C), 50 mM KCl, 1.5 mM MgCl2, 200 μM dNTPs, 2.5 U Taq DNA polymerase. The mix was subjected to 25 amplification cycles.Activity in reaction buffer of Pwo SuperYield DNA Polymerase PCR Mix is 100%; Pwo SuperYield DNA Polymerase PCR Mix is not available in U.S.
Application:
In PCR-mediated mutagenesis, DpnI nuclease has been used for the digestion of wild type DNA after PCR.
General description:
Dpn I requires methylation of adenine residues for activity and thus digests only GmATC sequences containing N6-methyladenine. Such methylated sequences are produced by dam methylase. This requirement for methylation distinguishes Dpn I from Mbo I, which is inhibited by dam methylation, and Sau3A I, which is not influenced by dam methylation. Dpn I also cleaves at a different position on the recognition sequence than Mbo I and Sau3A I.
Other Notes:
For life science research only. Not for use in diagnostic procedures.
Quality:
Absence of nonspecific endonuclease activities
1μg pBR322 DNA is incubated for 16 hours in 50μl SuRE/Cut Buffer A with an excess of Dpn I. The number of enzyme units which do not change the enzyme-specific pattern is stated in the certificate of analysis.
Absence of exonuclease activity
Approximately 5μg [3H] labeled calf thymus DNA are incubated with 3μl Dpn I for 4 hours at +37°C in a total volume of 100μl 50mM Tris-HCl, 10mM MgCl2, 1mM Dithioerythritol, pH approximately 7.5. Under these conditions, no release of radioactivity is detectable, as stated in the certificate of analysis.
Compatibility:
Dpn I generates fragments with blunt ends that are compatible with any blunt end.
DNA Profile:
Number of cleavage sites on different DNAs
• λ: 116
• φX174: 0
• Ad2: 87
• M13mp7: 8
• pBR322: 22
• pBR328: 27
• pUC18: 15
• SV40: 8
Specificity:
Recognition sites: GmAT*C
GmAT*C
Restriction site: GmA↓T*C
GmA↓T*C
Heat inactivation: No inactivation of Dpn I after incubation at 65 °C for 15 minutes.