Selecting an appropriate thermal cycler, also called a thermocycler or PCR machine, is crucial for the triumph of your PCR experiments. These experiments can be used for numerous applications, such as sequencing, genotyping, cloning, and mutagenesis.
Thermal cyclers, which are also referred to as PCR machines, thermocyclers, and DNA amplifiers, play a crucial role in amplifying and cloning DNA segments through the polymerase chain reaction (PCR) technique. These devices employ a thermal block that contains holes for accommodating the reaction mixtures in tubes. By utilizing a programmed set of timers, the head of the PCR system raises and lowers the temperature of the block, allowing the user to monitor the specific time and temperature required to achieve the desired reaction. Upon completion of the process, the results are analyzed using gel electrophoresis.
These instruments employ discrete, pre-programmed temperature steps to raise and lower the temperature of the samples secured within a holding block. This controlled temperature cycling facilitates double-strand denaturation, annealing, and polymerization processes. The amplified genetic material obtained can be utilized in various downstream applications, including cloning, sequencing, expression analysis, and genotyping. Researchers in the field of biology now have the luxury of choosing from a wide range of thermocycler models, each offering its unique advantages and benefits.
Thermal cyclers have been designed to effectively manage heat transfer and provide precise temperature control. These instruments offer complete programmability for each stage of a PCR protocol, allowing multiple protocols to be stored for future use. Additional software features may include user controls to restrict protocol editing, automatic report generation, and remote monitoring capabilities. The main components of a typical thermal cycler model comprise a sample block made of a thermally-conductive metal such as aluminum or silver, Peltier devices (also known as thermoelectric modules) for rapid heating and cooling functions, and a heated lid that prevents sample evaporation. Notably, the utilization of thermoelectric modules plays a crucial role in creating a self-contained instrument capable of fulfilling both heating and refrigeration requirements. A thermoelectric module functions as a solid-state device and operates as a heat pump. It consists of a thin ceramic wafer sandwiched between semiconductor blocks, such as bismuth telluride. When an electric current is applied, heat is transferred to either side of the device based on the current's direction, facilitating both the heating and cooling of samples. Any excess heat is dissipated using a heat sink and appropriate ventilation.
When considering buying a thermal cycler, there are a few things to keep in mind.
Firstly, temperature change is important. Most thermal cyclers regulate the temperature control of the block through a Peltier element. Ideally, the thermal cycler should instantly change the temperature inside the reaction tubes. However, in reality, it changes the temperature surrounding the outside of the sample tube. To ensure that reactions work properly, the temperature needs to be changed at precise times and as quickly as possible. The rate of temperature change required depends on your specific experimental needs. Every thermal cycler manufacturer aims to create heating blocks that change temperature evenly and as fast as possible. If rapid temperature changes are necessary for your work, look for thermal cyclers with a silver heating block. Silver heating blocks are ideal for quick temperature changes and promoting temperature uniformity throughout the heating block since silver conducts heat better than other materials used for heating blocks.
The thermal cycler block is available in a range of sample capacities, accommodating up to 384 sample tubes, for instance. The container type you choose for individual PCR reactions is related to the capacity issue. The most commonly used PCR reaction vessel is the 0.2-mL PCR tube. However, thermal cycler blocks that can hold various other containers, such as 0.5-mL PCR tubes, larger conical tubes, multiwell plates, and tube strips, can also be found.
Researchers used to apply a layer of mineral oil to each PCR reaction to prevent the reaction fluid from evaporating and condensing on the inside of the tube lids. Nowadays, instead of using mineral oil, many thermal cyclers feature a heated lid that lies on top of the samples in the temperature block, preventing evaporation and resulting condensation. This is not just for the sake of convenience, as evaporation and condensation can have catastrophic effects on data, especially in small-volume samples, by altering the concentration of reagents in PCR tubes and consequently resulting in skewed results or unwanted variation. A heated thermal cycler lid that is specialized can help to prevent such issues, particularly for sample tubes situated at the edges and corners of the heating block, which are more susceptible to evaporation and condensation.
Many older thermal cycler models and some of the high-quality models available today have a heating block that is one solid component. Nowadays, however, many manufacturers provide thermal cyclers that come with interchangeable heating blocks. This feature is particularly useful when you need to use a different kind of reaction vessel or a different number of samples. For example, you can easily switch from PCR tubes to multiple-well plates. Some manufacturers also offer thermal cyclers with modular heating block systems, where the single large heating block can be replaced by two or more smaller heating blocks that are the same size as the original. In some thermal cyclers, the smaller heating blocks can be controlled independently, allowing researchers to run multiple experiments at once. For instance, one researcher could be running three different experiments simultaneously, or three different researchers could each run their PCR reactions at the same time.
The PCR technique finds multiple applications in various fields, ranging from basic science to applied science. PCR Thermocyclers can arrest DNA fragments and beyond that, we can list different areas where PCR finds its applications. These areas include research, where PCR helps in cloning a segment of DNA. In medicine, PCR serves as a diagnostic tool for genetic diagnosis, genotyping, species detection, and disease detection. In paleontology and biological anthropology, PCR helps recover DNA. In forensic sciences, PCR helps establish a filial relationship between DNA samples. In agronomy and diversity, PCR allows the production of genetic traces of groups of crops of interest.