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BfiI (BmrI)

5'-A C T G G G (N)5^-3' 3'-T G A C C C (N)4^-5'		digested lambda DNA 0.7% agarose
#ER1591 Supplied with: 10X Buffer Tango™ 3X BfiI Stop Solution	50 u   1 ml 0.5 ml
#ER1592 Supplied with: 10X Buffer Tango™ 3X BfiI Stop Solution	250 u   1 ml 0.5 ml
Commercial Isoschizomers: BmrI, BmuI
Related Documents (in pdf, ~55 KB): Certificate of Analysis: #ER1591, #ER1592 MSDS (English) MSDS (English-USA) MSDS (German) Chart (in pdf, 721 KB) Brochure (in pdf, 316 KB)

Concentration
2 u/µl

Conditions for 100% Activity

Recommended buffer	Incubation temperature	Units for  overnight  incubation, u/µg DNA	Thermal inactivation, in 20 min	Enzyme activity, %
				B  (blue)	G  (green)	O  (orange)	R  (red)	Tango™  (yellow)
				1X	1X	1X	1X	1X	2X
Buffer Tango™	37°C	1.0	65°C	20-50	20-50	0-20	0-20	100	0-20

Termination of Digestion Reaction
Do not attempt to stop a digestion reaction by adding EDTA (see Note). If no further enzymatic manipulations with the digested DNA are required, use Stop Solution containing SDS or BfiI Stop Solution to terminate a reaction by incubating at 65°C for 10 min. If the digested DNA is to be used in down-stream manipulations, inactivate BfiI by incubation at 65°C for 20 min.

3X BfiI Stop Solution
(Warm up, to ensure the solution is homogeneous before use)
0.6% SDS, 0.05% bromophenol blue and 50% glycerol.

Storage Buffer
10 mM Tris-HCl (pH 7.5 at 25°C), 100 mM KCl, 1 mM DTT, 0.1 mM EDTA, 0.2 mg/ml BSA and 50% glycerol.

Ligation and Recleavage
After 10-fold overdigestion with BfiI, more than 40% of the digested pBR322 DNA fragments can be ligated and more than 90% of these can be recut.

Methylation Effects
Dam: Dcm: CpG: EcoKI: EcoBI:	never overlaps  never overlaps  never overlaps  may overlap  may overlap	- no effect. - no effect. - no effect. - effect not determined. - effect not determined.

Note

BfiI is the only known restriction enzyme that cleaves DNA specifically both in the presence and absence of Mg2+ ions in the reaction mixture (Sapranauskas, R. et al., J.Biol.Chem., 275, 30878-30885, 2000). Chelating of Mg2+ ions by EDTA does not inhibit BfiI activity and may cause non-specific products. The non-specific cleavage increases at temperatures higher than 37°C. Star activity is observed at a greater than 40-fold overdigestion with BfiI.

Double Digestions using Tango™ Buffer, DoubleDigest™

Optimal buffer	Enzyme activity, %
	Tango™  (yellow)
	1X	2X
Buffer Tango™	100	NR

NR: buffer is not recommended, since enzyme activity is less than 20%.

Number of Recognition Sites in DNA Molecules

Lambda	PhiX174	M13mp18/19	pBR322	pUC18/19	pUC57	pTZ19R/U	pBluescriptIIKS(-/+)	pBluescriptIISK(-/+)	pACYC177	pACYC184
4	0	1	5	2	2	2	2	2	1	3

Updated kovo 05, 2008 16:44