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Source
: Anabaena variabilis ATCC 27893
Compatible ends
(5'…GWC)
Bme18 I, Cpo I, Csp I, Eco47 I, PpuM I, Psp5 II, PspPP I, Rsr II, Rsr2 I, SanD I, Sin, VpaK11B I
Isoschizomers
: Bme18 I, Eco47 I, Sin I, VpaK11B I
Methylation Sensitivity
: Not sensitive to dam methylation
: Blocked by overlapping dcm methylation
: Blocked by overlapping CpG methylation
Unit definition :
One Unit is defined as the amount of Ava II 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 '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 Ava II and 42~43 ㎕ of sterile water in 50 ㎕ reaction mixture. Incubate at 37℃.
Heat Inactivation
: 65℃ for 20 min.
Activity in BeamsBio™ Buffer System
|
Rxn Buffer |
A |
B |
C |
D |
|
Activity(%) |
50~75 |
50~75 |
75~100 |
10~50 |
|
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 Ava II. And then incubate at 37℃ 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
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pBR322
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M13mp18
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SV40
|
|
35
|
73
|
2
|
1
|
8
|
1
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6
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Quality control
Overdigestion: The same band pattern as a digestion in one unit of Ava II for one hour was showing after incubation of 1 ㎍ of λ DNA with units of Ava II 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 Ava II.
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References
Murray, K., Hughes, S.G., Brown, J.S., Bruce, S.A., Biochem. J., 159: 317-322 (1976)/ Sutcliffe, J. G., Church, G. M., Nucleic Acids Res., 5: 2313-2319 (1978)/ Fuchs, C., Rosenvld, E. C., Honigman, A., Szybalsky, W., Gene, 4: 1-23 (1978)
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