Hot Start PCR
“Hot start” is a term used to describe the inactivation of a DNA polymerase until the initial denaturation step of PCR cycling. Hot start eliminates spurious amplification products resulting from non-specific priming events during reaction setup and initiation, and increases overall reaction efficiency. There are several methods used to inactivate DNA polymerases that include chemical modification (e.g. anhydrides or formaldehydes), physical modification (e.g. wax beads), aptamer binding, primer sequestration, antibody binding, and the addition of thermolabile blocking groups on dNTPs or primers.
Kapa Biosystems portfolio of DNA polymerases are offered in both standard and hot start formulations. Our hot start technology uses proprietary antibodies to inactivate the DNA polymerase until the initial denaturation step. Antibodies are denatured within the first 30 seconds of the initial denaturation at 95 ºC, providing a fast and efficient hot start method for high performance PCR. Hot start polymerases are recommended for:
- High-throughput PCR
- Amplification of complex targets susceptible to non-specific and mispriming
- Prolonged reaction setup on the bench or liquid handling
KAPA2G Fast HotStart DNA Polymerase offers faster and more efficient Hot Start PCR than competitor enzymes based on wild-type Taq. KAPA2G Fast HotStart DNA Polymerase is an engineered DNA polymerase designed specifically for high-throughput, high performance Fast PCR.
KAPA2G Robust HotStart DNA Polymerase is a novel DNA polymerase derived from wild-type Taq through molecular evolution, and is specifically designed for high performance PCR in chemically complex environments. The enzyme is resistant to many inhibitors commonly found in crude samples and is ideally suited for the routine amplification of difficult templates (i.e. GC-rich) and samples. The increased processivity and specific activity of KAPA2G Robust HotStart allows for improved PCR success rates across diverse primer sets and sample types.
KAPA HiFi HotStart DNA Polymerase is recommended for all applications where high fidelity is important or the target length is >3 kb (e.g. cloning, site-directed mutagenesis, resequencing of pooled amplicons). KAPA HiFi HotStart exhibits industry leading fidelity (100x higher fidelity than Taq polymerase), sensitivity, robustness, and an ability to amplify long and complex templates.
KAPA Taq HotStart DNA Polymerase is a wild-type Taq polymerase and engineered buffer system recommended for routine PCR applications.
KAPA Long Range HotStart DNA Polymerase is a blend of KAPA Taq DNA polymerase and the novel, engineered KAPA HiFi B-family DNA polymerase and is recommended for the amplification of long templates (up to 25 kb) or applications requiring improved senstivity. The fidelity of KAPA Long Range HotStart is 3x – 6x improvement over Taq polymerase.