Thermocyclers are integral to life science research. In molecular biology they are used for DNA sequencing, cloning, generation of probes, quantification of DNA and RNA, studying patterns of gene expression, detection of sequence-tagged sites, and many more techniques.
Thermocycler use extends beyond simple PCR and the amplification of nucleic acids to include both random and site-directed mutagenesis as well as the in vitro construction of recombinant DNA sequences. The fine temperature control and ability to hold a precisely set temperature with little fluctuation is also applicable to other techniques in which high-accuracy temperature control increases reproducibility between experiments. A thermocycler can be used in protocols where very strict temperature control is required such as heating tissue slides for in situ hybridization. The controlled temperature ramping provided by a thermocycler enables the study of temperature-dependent kinetics; for example, in addition to optimizing a PCR protocol, a thermocycler with a gradient feature allows for the determination of the optimum temperature for any enzymatic activity.
Several PCR machines and real-time PCR systems are available for educational use. All thermocyclers are fully functional standard machines. Using these PCR machines, students will gain a real research experience and an understanding of how PCR is being used in everyday testing.
Inquiry-based science teaching helps develop critical thinking skills. Integration of a memorable hands-on approach into the curriculum demonstrates practical applications of science, helps with retaining information, teaches how to interpret data, and keeps students engaged.
For life science education, a thermocycler is a good teaching tool for inquiry-based science. While learning how PCR analysis is used in practical ways, students are exposed to a range of scientific principles. Students can learn how PCR is used in fields such as forensics, diagnostics, animal conservation, and archaeology. A number of fun PCR amplification kits are available for teaching real-world PCR applications, for example, the Crime Scene Investigator PCR Basics™ kit and the Fish DNA Barcoding kit.
Speed is an important consideration for testing foods for pathogens. Many foodstuffs are perishable, so testing has to be fairly rapid or the food will not have a sufficient shelf life after testing is completed. For highly virulent pathogens such as E. coli O157:H7, speed and sensitivity is critical. This pathogen can be detected using a PCR kit in as little as 12 hours.
PCR is both more accurate and rapid than ELISA. Increased sensitivity means that an enrichment step is less likely to be required prior to assay. This shortens the overall time to getting a result and minimizes required hands-on time.
Use of automated PCR for pathogen testing, in addition to increasing speed, provides a higher throughput, lowers contamination, reduces the level of false positives, and decreases interlab variability. Walk-away automation provides automated DNA extraction and PCR setup, combined with automated data collection and analysis, bar codes for increased traceability, and LIMS integration.
Further, PCR kits are validated by several certification bodies. Validation studies have shown that the PCR kits are equivalent to, or better than, the various reference methods evaluated while providing results in much less time.
Culturing and ELISA were the main assays for testing for major food pathogens for many years. More recently, PCR assays and the ability to fully integrate a thermocycler into automated workflow have made pathogen testing faster and more sensitive, with a higher throughput.
There is an increasing array of types and uses of polymerase chain reaction, from reverse transcriptase PCR (RT-PCR) to Droplet Digital™ PCR. Which thermocycler, or thermocyclers, you choose depends on current and anticipated future needs. Considerations when choosing a thermocycler include the following:
One of the most difficult issues when buying a new PCR machine is deciding what your future needs will be. For instance, will there be a need for detection of low abundance targets? If so, Droplet Digital™ PCR might be a good option. If it is anticipated that there will be many different targets, a thermocycler with thermal gradient capability, providing rapid optimization of annealing temperatures, would be preferred