Store at -20℃. For Research Use Only!

CpG

Acc  I

Fbl I, Xmi I

Supplied with : 10X Reaction Buffer 'Acc I'

Concentration :            units/㎕

Lot # :                

Expiration date :                

Source : Acinetobacter calcoaceticus ATCC 49823

Compatible ends (5'…MK)

: Aci I, Acl I, BsaH I, BspD I, BstB I, HinP1 I, Cla I, Hpa II, HpyCH4 Ⅳ, Msp I, Nar I, Taq I, HinP1 I, CviA II, Hpy188 Ⅲ

Isoschizomers : Fbl I, Xmi I

Methylation Sensitivity

: Not sensitive to dam and dcm methylation

: Blocked by overlapping CpG methylation

Unit definition : One Unit is defined as the amount of Acc I required to completely digest 1 ㎍ of λ DNA in one hour at 37℃ in 50 ㎕ assay mixture.

Typical Reaction Conditions : Add 5 ㎕ of 10X Reaction Buffer 'Acc I'[final conc.: 50 mM Tris-HCl (pH 8.2 @ 37℃), 5 mM MgCl₂], 1~2 ㎕ of DNA(0.5~1.0 ㎍/㎕), 1 ㎕ of Acc I and 42~43 ㎕ of sterile water in 50 ㎕ reaction mixture. Incubate at 37℃.

Heat Inactivation : 80℃ for 20 min.

Activity in BeamsBio™ Buffer System

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

Activity in a complete PCR Mix : less than 10%
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 Acc I. And then incubate at 37℃ for one hour. Relative activity is determined by gel electrophoresis.

Storage Conditions : 10 mM Tris-HCl(pH7.4), 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
9	17	1	2	2	1	1

Quality control

Overdigestion: The same band pattern as a digestion in one unit of Acc I for one hour was showing after incubation of 1 ㎍ of λ DNA with         units of Acc I for 16 hr in 50 ㎕ of reaction mixture at 37℃ 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 Acc I.

Notes

- Acc I requires at least 13 base pairs beyond the end of its recognition sequence to cleave efficiently.

- The polylinkers found in M13 and pUC cloning vectors contain a single Acc I site.

- One unit of Acc I cleaves approximately 98% of pBR322 DNA.

- Activity retained after 10 minutes at 65℃. Five-fold enhanced activity occurs at 55℃.

References

Roberts, R.J. Nucleic Acids Res. 12: 167-204 (1984)/ Kang, S.C., Yoo, O.J., Korean J. Micorbiol. 23:13-19 (1985)/

Kawakami, B. et al., Agric. Biol. Chem. 55:1553-1559 (1991)

Beams Biotechnology Co., Ltd.

<Doc. Rev. 2008-10-15>