IsoFast® Bst 聚合酶

产品特点​

• 具有链置换性的 5'→3' DNA 聚合酶活性

• 缺乏 5'→3' 外切核酸酶活性

• 可在恒定温度下进行 DNA 合成

• 作用温度范围宽，最适温度为 65°C

• 对多种模板均可实现快速且稳定的扩增

• 配备先进的双组分缓冲系统，可在困难条件下获得更高产量

• 30 分钟快速流程

• 提供含荧光染料与不含染料的灵活规格

• 亦可提供 2x 预混液形式，以及用于 RNA 扩增的双酶一步法系统

• 甘油游离酶制剂

​获取报价与样品​

IsoFast® Bst 聚合酶是一种在大肠杆菌中表达的重组蛋白，代表嗜热脂肪地芽孢杆菌（先前称为嗜热脂肪芽孢杆菌）DNA 聚合酶的大片段。该蛋白部分催化 DNA 的 5'→3' 合成并具有链置换活性，但不包含 5'→3' 外切核酸酶结构域。[1]

​强大的链置换能力​

链置换是指酶能够解离下游遇到的双链模板 DNA 的氢键， essentially 在合成互补链的同时"解开"DNA 双链。IsoFast® Bst 聚合酶显示出强大的链置换活性，适用于全基因组扩增、多重置换扩增和等温扩增等方法。DNA 合成在恒定温度下进行，我们建议在 65°C 下运行反应。然而，IsoFast® Bst 聚合酶在较宽的温度范围（55°C 至 70°C，见图 4）内均表现良好，这为优化引物退火和链置换条件提供了更宽的范围。该酶在 80°C 下可被热灭活。

​快速且稳定的结果​

IsoFast® Bst 聚合酶专为快速扩增速度而设计，可在不同靶序列上获得快速且一致的结果（见图 1、2 和 3）。该酶配有先进的双组分缓冲系统，即使在困难条件下也能确保高产量和性能。可通过多种方法检测扩增，包括实时荧光检测和终点可视化。

​规格灵活​

为方便起见，该酶也可提供独立的荧光染料（可与任何 qPCR 仪兼容进行实时检测），以及 2x 预混液形式，以满足希望缩短准备时间的用户需求。无论选择何种规格，该酶均能提供一致的结果（见图 5）。

本产品不适用于 PCR。

1 Mead DA, McClary JA, Luckey JA, Kostichka AJ, Witney FR, Smith LM. Bst DNA polymerase permits rapid sequence analysis from nanogram amounts of template. Biotechniques. 1991 Jul;11(1):76-8, 80, 82-87.

​应用​

• 全基因组扩增

• 多重置换扩增

• 等温扩增

• 环介导等温扩增

• 分子诊断

1.Fast and consistent isothermal amplification performance: Isothermal amplification of scaffolding protein gene from M13mp18 ssDNA genome using IsoFast™ Bst Mix, NEB WarmStart® LAMP Kit, NEB Bst 2.0 DNA Polymerase and Thermo Bsm DNA Polymerase. The manufacturers' protocols were followed to set up the reaction mix. A primer mix consisting of 0.2μM for F3 and B3 primers, 1.6μM for FIP and BIP primers and 0.8μM for LoopF and LoopB primers was used. 8 serial dilutions of ssDNA template were used, corresponding to the number of copies of M13 genome indicated. The reaction was run at 65℃ for 100 minutes. A BioRad CFX96 Touch instrument was used to record fluorescence every 10 seconds. The time to result is the time required to reach the same fluorescent threshold. IsoFast™ Bst Mix shows fast amplification and consistent performance when compared to leading manufacturers.

2.Fast and consistent isothermal amplification performance: Isothermal amplification of BRCA1 gene from human genome (HeLa cells) using IsoFast™ Bst Mix, NEB WarmStart® LAMP Kit and NEB Bst 2.0 DNA Polymerase. The manufacturers' protocols were followed to set up the reaction mix. A primer mix consisting of 0.2μM for F3 and B3 primers, 1.6μM for FIP and BIP primers and 0.8μM for LoopF and LoopB primers was used. The total reaction volume was 25μL. 7 serial dilutions of ssDNA template were used, corresponding to the number of copies of human genome indicated. The reaction was run at 65℃ for 100 minutes. A BioRad CFX96 Touch instrument was used to record fluorescence every 10 seconds. The time to result is the time required to reach the same fluorescent threshold. IsoFast™ Bst Mix shows fast amplification and consistent performance when compared to leading manufacturers.

3.Fast and consistent isothermal amplification performance: Isothermal amplification of 16S gene from S. aureus genome using using IsoFast™ Bst Mix, NEB WarmStart® LAMP Kit and NEB Bst 2.0 DNA Polymerase. The manufacturers' protocols were followed to set up the reaction mix. A primer mix consisting of 0.2μM for F3 and B3 primers, 1.6μM for FIP and BIP primers and 0.8μM for LoopF and LoopB primers was used. The total reaction volume was 25μL. 7 serial dilutions of ssDNA template were used, corresponding to the number of copies of S. aureus genome indicated. The reaction was run at 65℃ for 100 minutes. A BioRad CFX96 Touch instrument was used to record fluorescence every 10 seconds. The time to result is the time required to reach the same fluorescent threshold. IsoFast™ Bst Mix shows fast amplification and consistent performance when compared to leading manufacturers.

4.Active over a broad temperature range from 55℃ to 70℃: Isothermal amplification of scaffolding protein gene (using M13mp18 ssDNA genome) was performed using IsoFast™ Bst Mix. The reaction was set up according to the protocol using a primer mix of 1.6μM for FIP and BIP primers and 0.8μM for LoopF and LoopB primers. The total reaction volume was 25μL. 6 serial dilutions of ssDNA template were used, corresponding to the indicated number of copies of the M13 genome. The reaction was run at 55.0℃, 57.4℃, 60.8℃, 65.2℃, 68.7℃, 70.1℃ for 34 minutes. A BioRad CFX96 Touch instrument was used to record fluorescence every 10 seconds. IsoFast™ Bst Mix is active over a broad temperature range.

5.Flexible IsoFast formats providing consistent results: Isothermal amplification of scaffolding protein gene (using M13mp18 ssDNA genome) was performed using IsoFast™ Bst Polymerase or IsoFast™ Bst Mix. The reactions were set up according to the protocol using a primer mix of 0.2μM for F3 and B3 primers, 1.6μM for FIP and BIP primers and 0.8μM for LoopF and LoopB primers. The total reaction volume was 25μL. 8 serial dilutions of ssDNA template were used, corresponding to the indicated number of copies of the M13 genome. The reaction was run at 65℃ for 34 minutes. A BioRad CFX96 Touch instrument was used to record fluorescence every 10 seconds. The time to result is the time required to reach the same fluorescent threshold. IsoFast™ Bst Polymerase gives consistent results as a standalone enzyme and buffer, and in a ready-to-use 2x mix format. Reactions 1000 x 25 μL Reactions