Thermo Fisher, EF0654, FastAP Thermosensitive Alkaline Phosphatase (1 U/μL)

Thermo Fisher, EF0654, FastAP Thermosensitive Alkaline Phosphatase (1 U/μL) Thermo Scientific FastAP Thermosensitive Alkaline Phosphatase catalyzes the release of 5'- and 3'-phosphate groups from DNA, RNA, and nucleotides. This enzyme also removes phosphate groups from proteins.
FastAP is a novel alkaline phosphatase, which is active in all Thermo Scientific restriction enzyme buffers as well as in PCR buffers. It dephosphorylates all types of DNA ends (blunt, 5'- and 3'-overhangs) in 10 minutes at 37°C. The enzyme is inactivated in 5 minutes at 75°C (seeFigure 1 in Supporting Data). Therefore, removal of alkaline phosphatase is not required prior to ligation.
Highlights
•Recombinant enzyme
•Fast dephosphorylation—10 minutes at 37°C
•Fast and complete inactivation—5 minutes at 75°C
•Simultaneous digestion and dephosphorylation of vector DNA
•100% activein restriction enzyme and PCR buffers
•PCR clean-up in conjunction with Exo I
•Protein dephosphorylationOne protocol for all types of DNA ends:
• 5'-overhangs
• 3'-overhangs
• blunt-ends
• single nucleotidesApplications
• Dephosphorylation of cloning vector DNA to prevent recircularization during ligation
• Simultaneous digestion and dephosphorylation of vector DNA
• PCR product clean-up: nucleotide degradation prior to sequencing of PCR product
• Dephosphorylation of nucleic acid 5'-termini prior to labeling withT4 Polynucleotide Kinase
• Other applications where dephosphorylation of DNA and RNA substrates is necessary
• Protein dephosphorylationNote
• Binding of FastAP Thermosensitive Alkaline Phosphatase to DNA may result in a band shift in agarose gels. To avoid this, incubate samples with6X DNA Loading Dye & SDS Solutionat 65°C for 10 minutes, and chill on ice prior to electrophoresis.
• FastAP Thermosensitive Alkaline Phosphatase is active in all restriction enzyme buffers and may be added directly to digested DNA. Heat inactivation of the restriction enzyme before dephosphorylation reaction is not necessary.
Compatible Buffer: PCR Buffer, Enzyme Buffer
Product Type: Alkaline Phosphatase
Quantity: 300 units
Concentration: 1 U/μL
Enzyme: Phosphatase
Unit Size: Each