85270-15-1

Basic information

• Product Name: SPH I
• Synonyms: APA I RESTRICTION ENZYME;APA I;SPH I;SPH I RESTRICTION ENZYME;RESTRICTION ENDONUCLEASES APA I;RESTRICTION ENDONUCLEASES SPH I;restriction endonuclease apa I from*acetobacter P;Nuclease, restriction endodeoxyribo-, SphI
• CAS NO: 85270-15-1
• Molecular Weight: 0
• EINECS: 286-569-8
• Product Categories: 3.1.x.x Acting on esters;3.x.x.x Hydrolases;Molecular Biology;Molecular Biology Enzymes;Molecular Biology Tested;Restriction EndonucleasesApplication Index;Restriction EnzymesEnzyme Class Index
• Mol File: 85270-15-1.mol
• Article Data: 0

Chemical Properties

• Appearance: /buffered aqueous glycerol solution
• Storage Temp.: −20°C
• CAS DataBase Reference: SPH I(CAS DataBase Reference)
• NIST Chemistry Reference: SPH I(85270-15-1)
• EPA Substance Registry System: SPH I(85270-15-1)

Safety Data

• WGK Germany: 1
• Hazardous Substances Data: 85270-15-1(Hazardous Substances Data)

Usage

General Description

Compatible endsApa I ends are not compatible with those generated by any other known restriction enzymes.IsoschizomersApa I is an isoschizomer to Bsp 120 I and Psp OMI.Methylation sensitivityApa I is inhibited by 5′-methylcytosine at the sites indicated (*) in the recognition sequence.Activity in SuRE/Cut Buffer SystemBuffer printed in bold face type is the buffer recommended for optimal activity:ABLMH100%10-25%50-75%50-75%0-10%Relative activity in complete PCR mixRelative activity in PCR mix (Taq DNA Polymerase buffer) is 100%. The PCR mix contained target DNA, primers, 10 mM Tris-HCl (pH 8.3, +20°C), 50 mM KCl, 1.5 mM MgCl2, 200 μM dNTPs, 2.5 U Taq DNA polymerase. The mix was subjected to 25 amplification cycles. Activity in reaction buffer of Pwo SuperYield DNA Polymerase PCR Mix is 10%. When supplemented with GC-RICH Solution activity is increased to >100%.Incubation temperature+30°CUnit definitionOne unit is the enzyme activity that completely cleaves 1 μg ? × Hind III fragments in 1 hour at +30°C in a total volume of 25 μl SuRE/Cut Buffer A.Heat inactivationThe enzyme can be heat inactivated by incubating it for 15 minutes at +65°C.Number of cleavage sites on different DNAsλAd2SV40φ X174M13mp7M13mp18pBR322pBR328pUC18112 1000000Ligation and recutting assayApa I fragments obtained by complete digestion of 1 μg λ × Hind III fragments are ligated with 1 U T4 DNA Ligase in a volume of 10 μl by incubation for 16 hours at +4°C in 66 mM Tris-HCl, 5 mM MgCl2, 5 mM Dithiothreitol, 1 mM ATP, pH 7.5 (at +20°C) resulting in >95% recovery of 1 μg λ × Hind III fragments.Subsequent re-cutting with Apa I yields >95% of the typical pattern of λ × Hind III× Apa I fragments.