phi29 DNA Polymerase
220.00 €+ excl. VAT
High purity, catalytically active phi29 overexpressed in E.coli and purified to single band homogeneity.
250 units, 1,000 units, 5,000 units
Single-pack, 2-pack, 3-pack
Not IATA restricted
Must be stored at -18°C to -20°C upon arrival
The phi29 DNA Polymerase is the only replicative polymerase required for the efficient replication of the Bacillus subtilis phage phi29 DNA. This is made possible by two properties of this enzyme 1) highly processivity without the requirements of any other replicative protein and 2) exceptional strand displacement coupled to the polymerization process. Therefore, the phi29 polymerase allows for very efficient isothermal DNA amplification. The enzyme is capable of amplification of up to 70 kb insertions per binding event and possesses a 3’→5’ proofreading exonuclease activity, acting preferentially on ssDNA or RNA. These properties make the phi29 polymerase an ideal tool to achieve isothermal strand displacement amplification. Specification
- VPCIR ™ phi29 holds marked processivity and strand displacement activity allowing for long DNA stretches to be synthesized (>70kb).
- High-fidelity polymerase – possesses a 3’-5’ exonuclease (proofreading) activity acting preferentially on ssDNA or RNA.
- High yields of amplified DNA even from small template amounts
- Isothermal polymerase – no need for thermal cycling.
- Due to heat lability of the enzyme, it is possible to use the amplification products directly in the downstream applications (PCR, restriction digestion, SNP genotyping, etc.).
The phi29 enzyme is supplied together with:
- Dilution Buffer: 50mM Tris-HCL pH 7.5; 1mM DTT; 50% glycerol; 0.5%Tween 20; 0.5% NP40; 500mM NaCl.
- Reaction Buffer 10x: 330 mM Tris-acetate; 100 mM Magnesium-acetate, 660 mM Potassium-acetate; 1% Tween20; 2mM DTT (to be added fresh).
- 5 M DTT
- Nucleases-free water
- Rolling circle amplification (RCA)
- Isothermal amplification for sequencing and cloning
- In situ genotyping with padlock probe
- Multiple displacement amplification (MDA)
- Unbiased whole genome amplification (WGA)
- Amplification of DNA for SNP and STR detection
- Cell-free amplification of DNA from single cells
- Pathogenic organisms or metagenomes
- Amplification of DNA from filter paper spot samples
- Amplification of DNA from environmental samples
- DNA template preparation for sequencing
- Protein-primed DNA amplification
- Recombination based-cloning
- Cell-free cloning of lethal DNA
- RNA-primed DNA amplification
Unit definition Phi29 unit has been determined by comparison with Thermofisher (EP0091) phi29 where 1 unit is defined as the amount of the enzyme that catalyzes the incorporation of 0.5 pmol of dCMP into a polynucleotide fraction in 10 min at 30 °C.
Recombinant GST-tagged phi29 enzyme is prepared by overexpressing the enzyme in E. coli followed by purification by affinity chromatography.
The enzyme is shipped cooled by gel packs and must be stored at -18°C to -20°C upon arrival. The temperature must be controlled to avoid that the enzyme preparation solidifies. This would destroy the enzyme integrity and cause a drop of the phi29 activity. The enzyme has a lifetime of 12 months when stored under optimal conditions.
Certificate of analysis of VPCIR ™ phi29 DNA Polymerase
1. Blanco, L.; Bernad, A.; Lázaro, J.M.; Martín, G.; Garmendia, C.; Salas, M. Highly Efficient DNA Synthesis by the Phage ϕ 29 DNA Polymerase. J. Biol. Chem. 1989, 264, 8935–8940, doi:10.1016/s0021-9258(18)81883-x.
2. Yokouchi, H.; Fukuoka, Y.; Mukoyama, D.; Calugay, R.; Takeyama, H.; Matsunaga, T. Whole-metagenome amplification of a microbial community associated with scleractinian coral by multiple displacement amplification using j 29 polymerase. 2006, 8, 1155–1163, doi:10.1111/j.1462-2920.2006.01005.x.
3. Larsson, C.; Koch, J.; Nygren, A.; Janssen, G.; Raap, A.K.; Landegren, U. In situ genotyping individual DNA molecules by target-primed rolling-circle amplification of padlock probes. 2004, 1, 1–6, doi:10.1038/NMETH723.
4. Salas, M.; Blanco, L.; Lázaro, J.M.; De Vega, M. The bacteriophage φ29 DNA polymerase. IUBMB Life 2008, 60, 82–85, doi:10.1002/iub.19.
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