Alkaline Phosphatase, TAB5

Catalog No. ON-179
Storage Condition -20℃
Source Recombinant E.coli
Purity ＞90% by SDS-PAGE

Alkaline Phosphatase, TAB5

Grade
Amount/Package: 1kU / centrifuge tube
SKU: ON-179-KU001-P
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Details

Product Information

Product description: Alkaline Phosphatase, TAB5 nonspecifically catalyzes the dephosphorylation of 5' and 3' ends of DNA and RNA phosphomonoesters. It also hydrolyses NTPs and dNTPs. It is completely inactivated by heating at 80℃ for 2 minutes.

Source: Recombinant E.coli

Concentration: 2.5U/μL

Unit Definition: One unit is defined as the amount of enzyme that will dephosphorylate 1μg of pUC19 vector DNA cut with a restriction enzyme generating 5' recessed ends in 30 minutes at 37℃. Dephosphorylation is defined as ＞ 95% inhibition of recircularization in a self-ligation reaction and is measured by transformation into DH5α.

Storage Buffer: 10mM Tris-HCl , 1mM MgCl2, 0.01mM ZnCl2, 50%(v/v) glycerol, pH7.4, 25℃
10X Phosphatase Reaction Buffer: 500mM Bis-Tris-Propane-HCl , 10mM MgCl2, 1mM ZnCl2, pH6, 25℃
10X IVT Reaction Buffer without Mg2+, Cat#ON-41, 400mM Tris PH7.9, 100mM DTT, 20mM Spermidine

Key points of operation


Protocol:

DNA as substance:

  1. Combine the follows on ice:
Component
Amount
5'-Phosphate-DNA
1μg
10X Phosphatase Reaction Buffer
2μL
Alkaline Phosphatase, TAB5
2.5-5 units
Nuclease-free H2O
to 20μL

  1. Incubate at 37℃, 30min
  2. (optional) Heat inactivity enzyme by incubating at 80℃, 2min.

RNA as substance:

  1. Heat Denature the RNA:
Component
Amount
5'-Phosphate-RNA
1μg
Nuclease-free H2O
to 15μL

  1. Incubate at 65℃, 10~15min, transfer the tube immediately to ice.
  2. On ice, combine the following reaction components in the order given:
Component
Amount
Heat denatured RNA from step 1 (above)
15μL
10X Phosphatase Reaction Buffer
2μL
Alkaline Phosphatase, TAB5
2.5-5 units
RNase Inhibitor (Recommend)
20 units
Total Reaction Volume
20μL

  1. Incubate at 37℃, 30min.
  2. Stop the reaction using any one of the following methods:
    a) immediate storage at –20℃ or –70℃.
    b) Add of EDTA to a final concentration of > 2mM.
    c) Phenol/chloroform extract then precipitate using salt/alcohol.

Security Information

Storage Conditions: -20℃

Quality Assurance: Free of endonuclease, exonuclease, nickase and RNase activities.

Physical Purity: ＞90% by SDS-PAGE.

Reference

1. Maria Rina, Charalambos Pozidis, et al. Eur. J. Biochem. 267, 1230-1238(2000).

2. Ellen Wang, Dimitris Koutsioulis, et al. J. Mol. Biol. 366, 1318-1331(2007).

3. Ping Li, Jon Beckwith, et al. Proc. Natl. Acad. Sci. USA.85,7685-7689(1988).

Specifications
  • Catalog No.
    ON-179
  • Source
    Recombinant E.coli
  • Purity
    ＞90% by SDS-PAGE
  • Storage Condition
    -20℃

Documentation
COA Contact Us

Product Information

Product description: Alkaline Phosphatase, TAB5 nonspecifically catalyzes the dephosphorylation of 5' and 3' ends of DNA and RNA phosphomonoesters. It also hydrolyses NTPs and dNTPs. It is completely inactivated by heating at 80℃ for 2 minutes.

Source: Recombinant E.coli

Concentration: 2.5U/μL

Unit Definition: One unit is defined as the amount of enzyme that will dephosphorylate 1μg of pUC19 vector DNA cut with a restriction enzyme generating 5' recessed ends in 30 minutes at 37℃. Dephosphorylation is defined as ＞ 95% inhibition of recircularization in a self-ligation reaction and is measured by transformation into DH5α.

Storage Buffer: 10mM Tris-HCl , 1mM MgCl2, 0.01mM ZnCl2, 50%(v/v) glycerol, pH7.4, 25℃
10X Phosphatase Reaction Buffer: 500mM Bis-Tris-Propane-HCl , 10mM MgCl2, 1mM ZnCl2, pH6, 25℃
10X IVT Reaction Buffer without Mg2+, Cat#ON-41, 400mM Tris PH7.9, 100mM DTT, 20mM Spermidine

Key points of operation


Protocol:

DNA as substance:

  1. Combine the follows on ice:
Component
Amount
5'-Phosphate-DNA
1μg
10X Phosphatase Reaction Buffer
2μL
Alkaline Phosphatase, TAB5
2.5-5 units
Nuclease-free H2O
to 20μL

  1. Incubate at 37℃, 30min
  2. (optional) Heat inactivity enzyme by incubating at 80℃, 2min.

RNA as substance:

  1. Heat Denature the RNA:
Component
Amount
5'-Phosphate-RNA
1μg
Nuclease-free H2O
to 15μL

  1. Incubate at 65℃, 10~15min, transfer the tube immediately to ice.
  2. On ice, combine the following reaction components in the order given:
Component
Amount
Heat denatured RNA from step 1 (above)
15μL
10X Phosphatase Reaction Buffer
2μL
Alkaline Phosphatase, TAB5
2.5-5 units
RNase Inhibitor (Recommend)
20 units
Total Reaction Volume
20μL

  1. Incubate at 37℃, 30min.
  2. Stop the reaction using any one of the following methods:
    a) immediate storage at –20℃ or –70℃.
    b) Add of EDTA to a final concentration of > 2mM.
    c) Phenol/chloroform extract then precipitate using salt/alcohol.

Security Information

Storage Conditions: -20℃

Quality Assurance: Free of endonuclease, exonuclease, nickase and RNase activities.

Physical Purity: ＞90% by SDS-PAGE.

Reference

1. Maria Rina, Charalambos Pozidis, et al. Eur. J. Biochem. 267, 1230-1238(2000).

2. Ellen Wang, Dimitris Koutsioulis, et al. J. Mol. Biol. 366, 1318-1331(2007).

3. Ping Li, Jon Beckwith, et al. Proc. Natl. Acad. Sci. USA.85,7685-7689(1988).

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