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Hph I
Recognition & cleavage sequence
Source: Haemophilus
parahaemolyticus
Reaction
conditions
1X
EzBuffer IV, 37℃
1X
FastCut Buffer, 37℃
FastCut
Buffer
Enzynomics restriction enzyme can cut substrate DNA in 5-15 min with FastCut Buffer
1X
EzBuffer IV
20
mM Tris-acetate (pH 7.9 @ 25℃), 50 mM
potassium acetate, 10 mM magnesium acetate, 100 μg/ml
BSA
Unit
definition
One
unit is defined as the amount of enzyme required for complete digestion of 1-μg bacteriophage λ at 37℃
for 1 hr in 50-μl reaction
mixtures.
Storage
10
mM Tris-HCl (pH 7.5 @ 25℃), 300mM NaCl,
0.1 mM EDTA, 1 mM dithiothreitol, 200 μg/ml
BSA, 50% glycerol.
Dilution
buffer: EzDiluent B
10
mM Tris-HCl (pH 7.4 @ 25℃), 300 mM
NaCl, 0.1 mM EDTA, 1 mM dithiothreitol, 500 μg/ml
BSA, 50% glycerol.
Heat
Inactivation
Hph
I can be inactivated at 65℃ for 20 min.
Methylation
sensitivity
dam methylation:
Conditionally sensitive
dcm methylation: Not
sensitive
CpG methylation:
Not sensitive
Prolonged
incubation
A
minimum amount of enzyme required to digest 1-μg substrate DNA for 16 hr; 0.25
U.
Relative
activity in EzBuffers
EzBuffer
I: 100%
EzBuffer
II: 75%
EzBuffer
III: 10%
EzBuffer
IV: 100%
FastCut
Buffer: 100%
Note
Activity
is inhibited by dam methylation partially overlapping its recognition
sequence.
● Material Safety Data Sheet
● Quality control
Overdigestion assay
DNA digested for 16 hr in 50-μl reaction
with 100U of enzyme resulted in same the DNA band patterns as those obtained
with 1 U of enzyme fo1 hr.
Ligation and recutting
More than 95% of DNA fragments (1 μg)
digestewith 50-fold excess enzyme can be ligated by T4DNA ligase (400 U) at 16℃ for 16 hr. Of the ligation products, > 95% can
be re-cut.
Extreme pure (EP)
No detectable
degradation of 32P-end labeled single-stranded and double-stranded
(5’-, 3’-overhang and blunt end) oligonucleotides occurred during incubation
with 100 U of enzyme for 4 hr.