Store at -20℃. For Research Use Only!

dam   ★   50℃

Bcl  I

Fba I, Ksp22 I

Supplied with : 10X Reaction Buffer 'C'

Concentration :            units/㎕

Lot # :                

Expiration date :                

Source : Bacillus  caldolyticus

Compatible ends (5'…GATC)

BamH I, BfuC I, Bgl II, Bsp143 I, BstMB I, BstX2 I, BstY I, Dpn II, Fba I, Ksp22 I, Kzo9 I, Mbo I, Mfl I, Nde II, Psu I, Sau3A I, Xho II

Isoschizomers

: Fba I, Ksp22 I

Methylation Sensitivity

: Blocked by overlapping dam methylation

: Not sensitive to dcm and CpG methylation

Unit definition : One Unit is defined as the amount of Bcl I required to completely digest 1 ㎍ of λ(dam^-) DNA in one hour at 50℃ in 50 ㎕ assay mixture.

Typical Reaction Conditions : Add 5 ㎕ of 10X Reaction Buffer 'C'[final conc.: 10 mM Tris-HCl(pH 7.9 @ 37℃), 10 mM MgCl₂, 50 mM NaCl, 1 mM DTT], 1~2 ㎕ of DNA(0.5~1.0 ㎍/㎕), 1 ㎕ of Bcl I and 42~43 ㎕ of sterile water in 50 ㎕ reaction mixture. Incubate at 50℃.

Heat Inactivation : No

Activity in BeamsBio™ Buffer System

Rxn Buffer	A	B	C	D
Activity(%)	10~50	75~100	75~100	50~75

Activity in a complete PCR Mix : 75~100%
Conditions: [10 mM Tris-HCl(pH 8.3 @ 25℃), 50 mM KCl, 1.5 mM MgCl₂, 2 pmol of primers, 200 μM dNTP Mix, 2.5 U Taq DNA polymerase and 1 ㎍ of DNA(template or substrate) in 50 ㎕ of reaction volume] After 30 amplication cycles, add 5 units of Bcl I. And then incubate at 50℃ for one hour. Relative activity is determined by gel electrophoresis.

Storage Conditions : 10 mM Tris-HCl(pH 7.4 @ 25℃), 50 mM KCl, 0.1 mM EDTA, 1 mM DTT, 200 ㎍/㎖ BSA, and 50% glycerol. Store at -20℃.

No. of Cleavage Sites

λ	Ad2	pUC18	φX174	pBR322	M13mp18	SV40
8	5	0	0	0	0	1

Quality control

Overdigestion: The same band pattern as a digestion in one unit of Bcl I for one hour was showing after incubation of 1 ㎍ of λ DNA with         units of Bcl I for 16 hr in 50 ㎕ of reaction mixture at 50℃ as determined by agarose gel electrophoresis.

Ligation-Recut: After a 10-fold digestion for one hour,        % of λ DNA fragments were ligated with T4 DNA ligase at 16℃ and        % of the ligated fragments could be recut with Bcl I.

Notes

- High glycerol concentration may result in star activity.

- Bcl I is not inactivated by heating and therefore should be extracted with phenol/chloroform and once with chloroform,

and then precipitate the DNA with ethanol.

References

Bingham, A.H.A., Atkinson, T., Sciaky, D., Roberts, R.J., Nucleic Acids Res. 5: 3457-3467 (1978)

Beams Biotechnology Co., Ltd.