ThermoSleuth™
Scans multiple primers against collections of genomes to find all hybridizations and amplicons.
ThermoSleuth™ finds all mishybridization sites, eliminates false positives in PCR
Solving Primer Specificity
ThermoSleuth™ automatically scans oligos against large genome databases to detect all thermodynamically stable hits. Different from BLAST, ThermoSleuth™ captures the important mishybridization hits by considering the following:
- Appropriate thermodynamic scoring, as BLAST doesn’t differentiate a GC pair from an AT pair
- Displays all false amplicons
- Consideration for 3’ extensibility and stable mismatches
- ThermoSleuth will score based on complementarity as opposed to similarity in BLAST
- ThermoSleuth allows for solution conditions, salt, buffers, additives and other experimental factors
The problem of false positives in PCR
The most common cause of false positives in PCR is the formation of false amplicons due to primer mishybridization which is not detected by BLAST because it scores hits incorrectly using sequence similarity rather than using the proper rules for match and mismatch complementarity. Additionally, many researchers do not have access to the computational capacity or storage required of modern genomic-based assay designs.
The solution to false positives in PCR
ThermoSleuth solves the problem of primer and probe mishybridization by scanning against whole genome databases such as the human genome or microbiome using the speed of BLAST while applying the proper thermodynamic model to mishybridization and crosshybridization results. The cloud integration of ThermoSleuth enables all researchers with the computational power to capture 50X as many thermodynamically stable and extensible hits as BLAST.
Pre-made and customizable genome collections
ThermoSleuth contains a huge repository of curated and pre-formatted sequence databases that can further be arranged into a customized sequence playlist. Some of the most popular playlists are shown below. Customers now have the capability to build a background database as large as they need without restricting memory, storage or computational capacity which is a huge source of pain for those researchers that have limited local computational capacity.
ThermoSleuth Features
| ThermoSleuth | Primer-BLAST | BLAST | |
|---|---|---|---|
| Single Genome Scan | ✓ | ✓ | ✓ |
| Multiple Genome Scan | ✓ | ||
| Playlist of Genomes | ✓ | ||
| Detects Extensible Hits | ✓ | ✓ | ✓ |
| Detects Probe Hybrdization | ✓ | ✓ | ✓ |
| Evaluates probe dyes and minor groove binders. | ✓ | ||
| Specificity Checking | ✓ | ✓ | |
| Sensitivity Checking | ✓ | ✓ | |
| Correctly scores mismatches by identity | ✓ | ||
| Detects Amplicons | ✓ | ✓ | |
| Adjustable primer extensibility settings | ✓ | ||
| Allows download of amplicon plus tails | ✓ | ||
| Graphic overview of results | ✓ | ✓ | |
| Genome view provides detailed thermodynamics | ✓ | ||
| Multiplex Capability | ✓ | ||
| Differentiates DNA from RNA backbones | ✓ | ||
| Modified Backbones | ✓ | ||
| Modified Bases | ✓ | ||
| Uses Monovalent Salt | ✓ | ||
| Uses Divalent Salt | ✓ | ||
| Simulates at different temperatures | ✓ | ||
| Simulates at different primer concentrations | ✓ | ||
| Correctly scores target dangling ends | ✓ | ||
| Correctly scores bulges | ✓ |