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Hin4I 

5'-^    8(N) G A Py (N)5 (A/C/G) T C (N)13-14^-3'
3'-^13-14(N) C T Pu (N)5 (T/G/C) A G (N)8    ^-5'

digested lambda DNA (dam-)
1.0% agarose
       
#ER1601
Supplied with:
10X Buffer Tango™
50X SAM		 50 u
 
1 ml
0.1 ml
#ER1602
Supplied with:
10X Buffer Tango™
50X SAM		 250 u
 
1 ml
2x0.1 ml
Commercial Isoschizomers: -

Related Documents (in pdf, ~55 KB):

Certificate of Analysis: #ER1601, #ER1602
MSDS (English)
MSDS (English-USA)
MSDS (German)
Chart (in pdf, 721 KB)
Brochure (in pdf, 316 KB)

Concentration
2 u/µl

Conditions for 100% Activity

Recommended
buffer		 Incubation
temperature		 Units for 
overnight 
incubation,
u/µg DNA		 Thermal
inactivation,
in 20 min

Enzyme activity, %
B 
(blue)		 G 
(green)		 O 
(orange)		 R 
(red)		 Tango™ 
(yellow)
1X 1X 1X 1X 1X 2X
Buffer Tango™ + 
0.05 mM S-adenosylmethionine		 37°C 0.2 65°C 20-50
(+SAM)		 20-50
(+SAM)		 0-20
(+SAM)		 0-20
(+SAM)		 100
(+SAM)		 0-20
(+SAM)

Storage Buffer
10 mM potassium phosphate (pH 7.4 at 25°C), 100 mM NaCl, 1 mM EDTA, 7 mM 2-mercaptoethanol, 0.2 mg/ml BSA and 50% glycerol.

Ligation and Recleavage
After 10-fold overdigestion with Hin4I, more than 95% of the digested DNA fragments can be ligated and only 50% of these can be recut due to the methylation of the recognition sequence by restriction enzyme.

Methylation Effects
Dam:
Dcm:
CpG:
EcoKI:
EcoBI:		  may overlap
 never overlaps
 may overlap
 never overlaps
 may overlap		 - blocked.
- no effect.
- cleavage impaired.
- no effect.
- effect not determined.

Note
  • Assayed using lambda DNA (dam-) (#SD0021).
  • Hin4I produces double-strand cuts on both sides from the interrupted recognition site. Its unique feature is a degenerate cleavage point on the 3' side of the recognition sequence (13 or 14 nt away).
  • Hin4I requires only Mg2+ for its activity, but is stimulated by S-adenosylmethionine. 0.05 mM S-adenosylmethionine gives more than a 10-fold increase in Hin4I activity. Still, a complete cleavage of some substrates with Hin4I is difficult to achieve.
  • Hin4I concentration is determined by a maximal cleavage level achieved when no change in the fragmentation patterns is observed with the further enzyme increase.

Double Digestions using Tango™ Buffer, DoubleDigest™

Optimal
buffer

Enzyme activity, %
Tango™ 
(yellow)
1X 2X
Buffer Tango™ + 
0.05 mM S-adenosylmethionine		 100
(+SAM)		 NR

NR: Buffer is not recommended, since enzyme activity is less than 20%.

Number of Recognition Sites in DNA Molecules

Lambda PhiX174 M13mp18/19 pBR322 pUC18/19 pUC57 pTZ19R/U pBluescriptIIKS(-/+) pBluescriptIISK(-/+) pACYC177 pACYC184
58 9 5 5 2 3 2 2 2 8 6

See also General Properties of Restriction Enzymes:

 

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Updated kovo 06, 2008 15:40