Ultra-Pure Phi29 DNA Polymerase

What it means: Purity Grade describes how thoroughly contaminating nucleic acid has been removed from the enzyme preparation. Ultra-pure here means physical, chemical and enzymatic purification steps, released against a non-specific DNase assay and a TaqMan qPCR DNA contamination test.

Why it matters:

• Phi29 amplifies whatever DNA is present, so any carried-over bacterial DNA in the enzyme is amplified alongside the sample.
• In whole-genome and single-cell amplification the input is minute, so even trace contamination can dominate the output.
• Release testing by qPCR gives a quantitative floor rather than a general purity claim.

How to interpret here: Ultra-pure is the property that makes this enzyme suitable for low-input MDA and WGA. Still run a no-template control on every batch — it is the only way to see contamination introduced by your own reagents.

Tip: Use dedicated pipettes and filter tips for isothermal amplification; the enzyme's sensitivity cuts both ways.

What it means: Operating Temp is the single temperature at which the reaction runs. Isothermal at 30°C means no thermal cycling is used — amplification proceeds continuously through strand displacement rather than heat denaturation.

Why it matters:

• A single temperature removes the need for a thermal cycler, so the reaction can run in a water bath, a heat block, or in the field.
• Because the template is never heated to 95°C, long products stay intact — this is why MDA yields amplicons tens of kilobases long.
• The enzyme is not thermostable; heating it will inactivate it rather than restart the reaction.

How to interpret here: Hold the reaction at 30°C for the full incubation, then heat-inactivate at the end. Do not attempt a cycling program.

Tip: Keep the reagents at −20°C except while in use, as the supplied buffer contains BSA.

What it means: Buffer Included describes the reaction buffer supplied with the enzyme. This product ships with 500 µl of 5X Ultra-Pure Phi29 Reaction Buffer, matched to the 200-reaction enzyme pack. The buffer contains 0.5 µg/µl BSA.

Why it matters:

• BSA stabilises the polymerase across the long isothermal incubation and reduces loss to tube walls in small reactions.
• Phi29 activity depends closely on salt and pH; a generic polymerase buffer will not reproduce the processivity it is bought for.
• A buffer containing protein is more perishable than a salt-only buffer, so handling matters more than usual.

How to interpret here: Dilute the 5X buffer five-fold in the final reaction. Because of the BSA, keep it at −20°C at all times except while pipetting.

Tip: Thaw on ice rather than at room temperature, and avoid repeated freeze-thaw of the buffer.