Polymerase chain reaction (PCR) uses enzymes to replicate a portion of DNA for analysis. Taq polymerase, extracted from a bacterium, is key. PCR requires six components and involves denaturation, annealing, and extension. PCR has various uses, including paternity testing and DNA analysis of extinct species or crime scenes.
Polymerase chain reaction (PCR) uses enzymes to replicate a portion of a deoxyribonucleic acid (DNA) strand en masse for easier analysis, such as searching for genes of interest. Like the nuclear chain reaction, the polymerase chain reaction is an exponential process that proceeds as long as the raw materials are available to sustain the reaction. In contrast to DNA replication in the natural world, PCR can only replicate small enough pieces of DNA, with an upper ceiling of about 2-3 kilo base pairs (kb). It uses inanimate enzymes to accomplish its replication effect, distinguishing it from other copying approaches that use active organisms.
A modern polymerase chain reaction requires six basic components to function: the segment of DNA to be copied, primers to delimit the segment, Taq polymerase to perform the copy, DNA nucleotides to serve as raw material, a chemical buffer environment, and a machine called thermal cyclist. The thermal cycler often contains multiple tubes with multiple PCRs, each containing 15 to 100 microliters, just under a cubic millimeter of water. About a hundred base nanograms of DNA are used.
Taq polymerase, the key ingredient for a polymerase chain reaction, is extracted from a deep-sea, hot spring-dwelling bacterium, Thermus aquaticus. It works well for copying, but not perfectly, making an error about once every 8 million base pairs. Prior to Taq polymerase, other polymerases were used, but many of them break down at the temperatures required for the reaction to begin. The heating cycle is complicated, but includes temperatures that briefly go nearly to the boiling point, so duration in the polymerase is of the essence.
The basic steps of PCR are as follows. All the ingredients are mixed together in a small vial, usually with a volume of 200 micrograms. The mixture is heated near boiling point to break the hydrogen bonds in double-stranded DNA, creating single strands that are amenable to copying. This is called denaturation. The longer the wire to be copied, the longer the denaturation process takes.
The next step of the polymerase chain reaction is called annealing. Primers, which are short, custom-made strands of DNA, are specifically designed to bind to sites at the beginning and end of the segment to be copied. If the primers are designed incorrectly or the temperature in this step is wrong, the primer will randomly bind to the DNA, resulting in the wrong segment being copied. Most primers melt at about two-thirds boiling point and annealing, a 1-2 minute process, occurs a few degrees below this.
The last steps of the PCR are called extension and final extension. This is where the magic happens. The polymerase copies the segment of DNA rapidly, making millions upon millions of copies in minutes. Usually, one cycle consists of all of the previous steps, repeated about twenty to thirty times.
The result is a bunch of copied DNA. Polymerase chain reactions have a variety of uses, including paternity testing, determining the presence or absence of a genetic defect or viral DNA, cloning a gene, introducing specific mutations, DNA analysis of extinct species or dead people , “genetic fingerprint” at the crime scene and more.
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