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HpaII, (HpaII)

Marka:EURx

Ürün Ölçü Fiyat TL Miktar
HpaII, (HpaII)
E2260-01
2 000 u 52,8 EUR
+%20 KDV
2.597,76 TL
HpaII, (HpaII)
E2260-02
10 000 u 208 EUR
+%20 KDV
10.233,60 TL

HpaII

(Haemophilus parainfluenzae)

5'-C C G G-3'
3'-G G C C-5'

Reaction Temperature: 37°C

Prototype: HpaII

Inactivation Temperature (20 min): 65°C

Reaction Buffer:

1x Low Buffer: 10 mM Tris-HCl (pH 7.0 at 37°C), 10 mM MgCl2, 1 mM dithiothreitol, 100 µg/ml bovine serum albumin.

Unit Definition: One unit is the amount of enzyme required to completely digest 1 µg of lambda DNA in 1 hr in a total reaction volume of 50 μl.

Assay Conditions: 10 mM Tris-HCl (pH 7.0 at 37°C), 10 mM MgCl2, 1 mM dithiothreitol, 100 μg/ml bovine serum albumin, 1 μg of lambda DNA. Incubation is at 37°C for 1 hr in a reaction volume of 50 µl.

Storage Buffer: 10 mM Tris-HCl (pH 7.5 at 22°C), 50 mM KCl, 0.1 mM EDTA, 1 mM dithiothreitol, 100 μg/ml bovine serum albumin and 50% (v/v) glycerol.

Storage Conditions: Store at –20°C.

Quality Control:

Non-specific Endonucleases: Incubation of 10 units of HpaII with 1 µg of lambda DNA at 37°C for 16 hrs (a 160-fold over-digestion) resulted in the same sharp characteristic banding pattern as the standard assay reaction, as determined by agarose gel electrophoresis.

3’-Exonuclease: 5, 10 and 20 units of HpaII and 0.15 µg (1.15 pmol of 3’-ends) of lambda/TaqI fragments (3’-labelled with T4 DNA Polymerase and [³H]dGTP and [³H]dCTP), incubated for 1 hr at 37°C resulted in 0.03-slope of %-end label released per unit of enzyme. Reaction volume 10 µl.

5’-Exonuclease/5’-Phosphatase: Incubation of 5, 10 and 20 units of HpaII with 0.12 µg (1.07 pmol of 5’-ends) of [5’-³²P] lambda/HaeIII fragments for 1 hr at 37°C resulted in 0.04-slope of %-end label released per unit of enzyme. Reaction volume 10 µl.

Ligation-recut Assay: 7.5 µg of lambda DNA was incubated with 30 units of HpaII for 1 hr at 37°C in a reaction volume of 150 µl. 1.1 µg was removed for an assay. The remaining DNA was incubated with 2 Weiss units of T4 DNA Ligase for 1 hr at 22°C. 3.2 µg were removed for an assay. The remainder of the DNA in the ligation mixture was recovered and then incubated with 50 units of Hpa II for 1 hr at 37°C. Aliquots of the cut, ligated and recut DNA were electrophoresed on an agarose gel. Results show >95% ligation and >95% recutting.

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