Store at -20℃. For Research Use Only!

★  65°C

Tth111  I

Asp I, PflF I, Psy I, Tel I

Supplied with : 10X Reaction Buffer 'B'

Concentration :            units/㎕

Lot # :                

Expiration date :                

Source : Thermus  thermophilus  111

Compatible ends (5'…N)

AclW I, Alw I, Asp I, Bcc I, Bin I, Bis I, Bme1390 I, BmgT120 I, BmrF I, BstEN I, EcoN I, Fnu4H I, Fsp4H I, Ita I, MspR9 I, PflF I, Ple I, Psy I, Pps I, Sat I, ScrF I, Tel I, Xag I

Isoschizomers : Asp I, PflF I, Psy I, Tel I

Methylation Sensitivity

: Not sensitive to dcm, dam and CpG methylation

Unit definition : One Unit is defined as the amount of Tth111 I required to completely digest 1 ㎍ of λ DNA in one hour at 65℃ in 50 ㎕ assay mixture.

Typical Reaction Conditions : Add 5 ㎕ of 10X Reaction Buffer 'B'[final conc.: 6 mM Tris-HCl(pH 7.5 @ 37℃), 6 mM MgCl₂, 50 mM NaCl, 1 mM DTT], 1~2 ㎕ of DNA(0.5~1.0 ㎍/㎕), 1 ㎕ of Tth111 I and 42~43 ㎕ of sterile water in 50 ㎕ reaction mixture. Incubate at 65℃.

Heat Inactivation : No

Activity in BeamsBio™ Buffer System

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

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 Tth111 I. And then incubate at 65℃ for one hour. Relative activity is determined by gel electrophoresis.

Storage Conditions : 10 mM Tris-HCl(pH 7.4 @ 25℃), 500 mM NaCl, 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
2	12	0	0	1	0	0

Quality control

Overdigestion: The same band pattern as a digestion in one unit of Tth111 I for one hour was showing after incubation of 1 ㎍ of λ DNA with         units of Tth111 I for 16 hr in 50 ㎕ of reaction mixture at 65℃ 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 Tth111 I.

Notes

- Conditions of low ionic strength, high enzyme concentration(>5 U/㎍ DNA), glycerol concentration > 5%, or pH > 8.0

may result in star activity.

- Tth111 I produces fragments that have a single-base 5´ extension which are more difficult to ligate than blunt-end.

- Tth111 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

Shinomiya, T., Sato, S. Nucleic Acids Res. 8: 43-56 (1980)

Beams Biotechnology Co., Ltd.