An integrated thermal cycler is a device used in molecular biology and biochemistry to perform polymerase chain reactions (PCR). It combines the functions of a thermal cycler and a PCR machine into one integrated system. This allows for the automated heating and cooling of PCR samples in a controlled manner, enabling the amplification of specific DNA sequences in a laboratory setting.
How is it used?
Integrated thermal cyclers are used in molecular biology and biochemistry for performing PCR reactions. PCR is a laboratory technique that allows for the amplification of specific DNA sequences. The integrated thermal cycler automates the process of heating and cooling the PCR samples, which is necessary for the success of the reaction. The PCR process typically involves the following steps: denaturation, annealing, and extension. The integrated thermal cycler can be programmed to perform these steps automatically, making the PCR process more efficient and consistent. The amplified DNA can then be used for various downstream applications, such as sequencing, genotyping, and gene expression analysis.
PCR requirements needed
Four parts (reagents and chemicals) are required to carry out the process of PCR:
- A DNA or an RNA sample (from blood, saliva scrapings of skin, hair or scraping, etc.)
- DNA primers are short single-stranded DNA that encourages the synthesis of a complementing nucleotide
- DNA polymerase is an enzyme that assists in synthesizing a complementary DNA strand. DNA
- Nucleotide solution mix that contains the amino acid adenine (A) and thymidine (T), Cytosine (C), and the amino acid guanine (G) that is used to construct duplicate DNA strands
PCR process steps
The process of PCR has four stages: collection, preparation, amplification, and post-PCR cleanup. The PCR machine’s steps take place during the amplification stage. It starts with a fragment of DNA placed into an appropriate tube, along with the reagents and other chemicals mentioned above. The tube is put into the PCR machine or thermal cycler. The thermal cycler moves your solution via a three-step denaturation, annealing, and extension process.
The steps of a thermal cycler typically include the following:
- Denaturation: The sample is heated to a high temperature (typically around 95-98°C) to separate the double-stranded DNA into single strands.
- Annealing: The sample is cooled to a lower temperature (typically around 50-60°C) to allow the complementary primers to anneal to the single-stranded DNA template.
- Extension: The sample is heated to a moderate temperature (typically around 72°C) to allow the Taq polymerase enzyme to extend the primers and create new DNA strands.
- Repeat: The denaturation, annealing, and extension steps are repeated for a specified number of cycles, usually between 20 and 40.
- Final extension: The sample is held at a high temperature (typically around 72°C) for a specified amount of time to ensure the complete extension of the DNA strands.
- Cooling: The sample is cooled to 4°C to end the reaction and stabilize the amplified DNA.
These steps are programmed into the thermal cycler, which automatically heats and cools the samples to the appropriate temperatures. Using a thermal cycler helps ensure consistent and efficient amplification of the target DNA sequences.