Direct PCR is a direct nucleic acid amplification technique using animal or plant tissues as template. In many ways, direct PCR works like standard PCR. However, the main difference is that the specialized buffer used in direct PCR can be directly used for PCR reaction without the need of nucleic acid extraction, which has high requirements for the tolerance of the enzyme and the compatibility of the buffer involved in direct PCR reaction .
Although PCR inhibitors are more or less present in common samples, direct PCR can still achieve reliable amplification under the action of optimized enzyme and buffer. While the standard PCR reaction requires high-quality nucleic acid as the template. If the template contains protein or other impurities, the PCR reaction will be inhibited. Therefore, with great convenience and robust amplification, direct PCR is becoming one of the most popular molecular biology techniques nowadays.
The earliest application of direct PCR is in the fields of animals and plants, such as the blood, tissue and hair of animals, or the leaves and seeds of plants, to study genotyping, transgenic, plasmid detection, gene knockout analysis, DNA source identification, species identification, SNP analysis and other fields. There are some common characteristics in these fields, that is, the content of target gene is relatively high, and nucleic acid extraction is troublesome. Therefore, direct PCR can not only save time, but also save cost. Direct PCR for pathogen detection is just developed in recent years, especially during the epidemic of COVID-19. Many researchers in both academia and industry have made great efforts in this direction.
At SBS Genetech, we are at the forefront of offering innovative solutions for direct PCR. One of the core enzymes is our PrimeTaq™ HotStart Direct PCR DNA Polymerase, which has hot start property and high tolerance to many PCR inhibitors. High tolerance delivers convenience for DNA amplification by allowing PCR directly from samples without prior DNA purification. With hot start property, the polymerase is 100% inactive below 50°C and can be completely recovered only after heating at 95°C for 5 min. Therefore, the system can effectively inhibit non-specific PCR amplification, greatly improving the specificity and sensitivity.
As direct PCR is not only used in the amplification of DNA template, but also widely used in the reverse transcription PCR of RNA template, we have also developed the enzyme for direct PCR of RNA template. Our PrimeTaq™ HS DP DNA/RNA Polymerase not only has hot start property and high tolerance to many PCR inhibitors like PrimeTaq™ HotStart Direct PCR DNA Polymerase, but also has strong RNA template dependent reverse transcriptase activity, which makes it ideal for the direct PCR of RNA template.
Direct PCR is also useful in real time PCR reaction, which requires the reaction system to have strong anti-interference ability of background fluorescence and antagonistic ability to endogenous fluorescence quenching agents. Both PrimeTaq™ HotStart Direct PCR DNA Polymerase and PrimeTaq™ HS DP DNA/RNA Polymerase have 5'- 3' exonuclease activity for TaqMan probe cleavage, and the optimized reaction system has strong anti-interference ability and antagonistic ability, so a robust real time PCR reaction is guaranteed.
With great convenience and efficiency, direct PCR is becoming progressively more important as time goes on. At SBS Genetech, we will keep developing novel tools for direct PCR, contributing to the scientific community with our best efforts.