In the intricate world of molecular biology techniques, DNA polymerases play a pivotal role, acting as molecular architects in the construction of genetic material. Among these, Bst polymerase and Taq polymerase stand out as two indispensable tools, each with its own set of characteristics and applications. While they may share similarities in function, a closer examination reveals distinctive features that make them suitable for different experimental needs.

Originating from different thermophilic bacteria, Bst polymerase and Taq polymerase boast unique biological pedigrees. Bst polymerase emerges as a large fragment of DNA polymerase I from Geobacillus stearothermophilus, a resilient bacterium thriving in extreme thermal environments. In contrast, Taq polymerase finds its roots in Thermus aquaticus, another thermophile known for its ability to endure scorching temperatures. This disparity in origin sets the stage for the subsequent differences in their properties and applications.

Thermostability serves as one of the most prominent distinctions between Bst polymerase and Taq polymerase. Bst polymerase exhibits a moderate level of thermal stability, making it an ideal candidate for isothermal DNA amplification methods. This characteristic allows researchers to conduct DNA amplification reactions at a constant temperature, simplifying experimental setups and reducing equipment requirements. Conversely, Taq polymerase boasts exceptional thermostability, capable of withstanding the rigorous temperature cycles of polymerase chain reactions (PCR) without succumbing to denaturation. This resilience in the face of extreme heat has cemented Taq polymerase's status as a cornerstone of PCR technology.

Another differentiating factor lies in the processivity of these enzymes. Processivity refers to the enzyme's ability to catalyze multiple reactions without dissociating from the template DNA. Bst polymerase has been noted for its superior processivity compared to Taq polymerase, enabling it to efficiently replicate long stretches of DNA with fewer interruptions. This attribute makes Bst polymerase particularly valuable in applications requiring the amplification of lengthy DNA fragments or complex genomic regions.

Fidelity, or the accuracy of DNA replication, also diverges between Bst polymerase and Taq polymerase. Over the years, Bst polymerase has undergone refinements aimed at enhancing its fidelity, salt tolerance, and other critical characteristics. These improvements have led to a reduction in error rates, bolstering the reliability of DNA amplification processes involving Bst polymerase. Conversely, Taq polymerase is notorious for its relatively high error rate, primarily attributed to the absence of a 3′–5′ exonuclease proofreading activity. Despite this drawback, Taq polymerase remains indispensable in PCR applications, where rapid amplification often takes precedence over absolute fidelity.

The applications of Bst polymerase and Taq polymerase reflect their unique properties and strengths in molecular biology research and diagnostics. Bst polymerase finds widespread use in isothermal DNA amplification methods such as Loop-Mediated Isothermal Amplification (LAMP), Whole Genome Amplification (WGA), and Rolling Circle Amplification (RCA). These techniques leverage Bst polymerase's strand displacement activity and moderate thermostability to achieve rapid and robust DNA amplification under isothermal conditions. On the other hand, Taq polymerase remains a staple in conventional PCR applications, where its exceptional thermostability and versatility make it indispensable for amplifying specific DNA targets with high precision and efficiency.

In conclusion, while Bst polymerase and Taq polymerase share the common role of DNA polymerases in molecular biology techniques, their distinct properties and origins give rise to specialized applications. Bst polymerase, with its moderate thermostability, superior processivity, and improved fidelity, excels in isothermal DNA amplification methods. Meanwhile, Taq polymerase's exceptional thermostability and versatility continue to make it the go-to choice for traditional PCR applications. Understanding these differences empowers researchers to select the most suitable enzyme for their specific experimental needs, ultimately advancing the frontiers of molecular biology research and diagnostics.

At SBS Genetech, we have pioneered the development of a diverse array of novel Bst polymerase variants meticulously tailored to amplify their utility in molecular biology research and diagnostics. Let's delve deeper into these groundbreaking solutions.

Bst DNA Polymerase Large Fragment: Cost-effective version for conventional strand displacement reaction

Bst DNA Polymerase Large Fragment  is the portion of the Bacillus stearothermophilus DNA Polymerase protein that contains the 5´ → 3´ polymerase activity, but lacks 5´ →3´ exonuclease activity. It has basic strand displacement activity and can be used for isothermal amplification.

However, without further genetic engineering, the amplification speed, impurity tolerance, and specificity of Bst DNA Polymerase (Large Fragment) are not ideal. So we don't recommend this enzyme for most of your experiments.

Bst DNA Polymerase: Ideal for isothermal amplification of DNA template

Bst DNA Polymerase is also derived from Bacillus stearothermophilus DNA polymerase I. Its 5 '- 3' exonuclease activity was removed by genetic engineering, while the 5 '- 3' polymerase activity was retained.

As the name suggests, Bst DNA Polymerase has a strong strand-displacement ability for DNA template, so it is an excellent enzyme for isothermal amplification.

Compared with wild-type Bst DNA polymerase (large fragment), Bst DNA Polymerase has been greatly improved in terms of amplification speed, yield, salt tolerance, and thermal stability.

At the same time, Bst DNA Polymerase can be amplified with dUTP as substrate, while Bst DNA Polymerase (Large Fragment) has no such activity.

Bst Polymerase: For isothermal amplification of both DNA and RNA template with a single enzyme system

Bst Polymerase is also derived from Bacillus stearothermophilus DNA polymerase I. Its 5'- 3' and 3'-5' exonuclease activity was removed by genetic engineering, while the 5'- 3' polymerase activity was retained.

Compared with wild-type Bst DNA polymerase (large fragment) and Bst DNA Polymerase, Bst Polymerase has better isothermal amplification activity and stronger reverse transcription activity. Single enzyme system reaction can be realized in the isothermal amplification experiment with RNA as a template.

Bst Polymerase has good reverse transcription activity at 60-65°C, which can effectively solve the reverse transcription of the RNA template with secondary complex structure, while Bst DNA Polymerase and wild-type Bst DNA polymerase (large fragment) do not have this activity.

Bst DNA/RNA Polymerase: Ideal for isothermal amplification of RNA template

Bst DNA/RNA Polymerase is a mixture of Bst Polymerase and extremely thermostable reverse transcriptase (65°C tolerant), which is suitable for the isothermal amplification reaction of RNA. It can detect low-sensitivity RNA molecules.

Bst DNA/RNA Polymerase is suitable for isothermal amplification reaction of both DNA and RNA templates, which can detect low-sensitivity nucleic acid templates with great efficiency and specificity. Besides, with a special preparation process, this enzyme has a fast amplification rate and high tolerance to impurity.

Since Bst DNA/RNA Polymerase is extremely thermostable and also provides sensitive reverse transcriptase activity, it is reported to have higher sensitivity at high Ct values.^[1]

Bst P DNA/RNA Polymerase: The most advanced enzyme solution

Bst P DNA/RNA Polymerase is an upgraded version of Bst DNA/RNA Polymerase through enzyme electronic re-structure and evolution screening (in silico Design & in vitro Evolution), which is generally used for LAMP or RT-LAMP amplification of DNA or RNA.

Performance improvements include:

• The whole Bst P DNA/RNA System includes hot start Aptamer, which ensures that the enzyme activity blocking efficiency is >95% at <30°C, and the enzyme activity is completely released within 1 min at >60°C. This characteristic facilitates the establishment of the reaction system at room temperature and greatly reduces the non-specific amplification at low temperatures.
• The reaction temperature is further raised to 70°C, which greatly reduces the formation of primer dimer, improves the amplification specificity, and makes the nucleic acid release of crude samples more sufficient.
• The whole portfolio contains Helicase, so Premium LAMP amplification (pLAMP) is allowed without using F3/B3 primers. At the same time, Helicase has the function of assisting in strand unwinding, which further reduces the concentration of FIP/BIP primers. This will further reduce non-specific amplification and greatly improve amplification homogeneity.

Comparison between the most common Bst DNA Polymerase and the latest Bst P DNA/RNA Polymerase

In conclusion, the selection of a Bst polymerase variant is intricately tied to research objectives, with each offering distinct capabilities and benefits. Our diverse array of Bst polymerases at SBS Genetech is meticulously curated to meet varied requirements.

Reference

[1] Lu S, Duplat D, Benitez-Bolivar P, León C, Villota SD, Veloz-Villavicencio E, et al. (2022) Multicenter international assessment of a SARS-CoV-2 RT-LAMP test for point of care clinical application. PLoS ONE 17(5): e0268340. https://doi.org/10.1371/journal.pone.0268340