The technique has revolutionized research to suchThe technique has revolutionized research to such

The development of the polymerase chain reaction (PCR) technique has been a major breakthrough in the scientific world. The technique has revolutionized research to such an extent that it can be considered as one of the major reasons for the boom the field has experienced over the last many years.INVENTION OF PCRKary Mullis is generally credited for inventing PCR in 1983 while working for Cetus Corporation in Emeryville, California. He was trying to think of a way to detect the point mutations using Sanger-type DNA sequencing, employing DNA polymerase in the presence of an oligonucleotide primer and dideoxynucleotides triphosphates (ddNTPs). However, he faced a problem that sequencing a single copy gene within the stretches of the human genome was impossible. Thereby, he needed a way to increase the concentration of the specific gene of interest.One day, while driving his Honda Civic on Highway 128 from San Francisco to Mendocino, an idea stuck his mind that by using a pair of primers to bracket the desired DNA sequence and performing multiple cycles of denaturation, annealing and polymerization he could exponentially amplify a small strand of DNA.Thus, the idea of PCR was born. He used E. coli DNA polymerase enzyme to amplify DNA after annealing of the primer but it got destroyed during the denaturation step so it had to be replenished after every cycle. Cetus workers overcame this problem by developing the first thermal cycler named “Mr Cycle”, which automatically added new polymerase after each heating step. In 1985, Mullis came up with the use of polymerase isolated from the extremophilic bacterium Thermophilus aquaticus. The polymerase, known as Taq polymerase, has optimal activity at 72°C and could withstand the 94°C required for denaturation of the DNA. This achievement along with advances in oligonucleotide synthesis made PCR cost effective and a convenient technique.APPLICATIONS OF PCRThe polymerase chain response is utilized by a wide range of researchers in a continually expanding scope of experimental orders.? Diagnosis of genetic diseasesPCR can be used to detect inherited genetic changes, even before birth or spontaneous genetic mutations. Examples include:• Genetic counselling – screening the parents for genetic disease before deciding on having children• Preimplantation diagnosis – screening for genetic disease before implantation of an embryo in IVF (in vitro fertilisation)• Screening for genetic disease before birth using tissue samples from the chorionic villus (the membranes found between the mother and unborn baby); foetal tissue from the amniotic fluid (the fluid around the unborn baby); or the small quantities of foetal DNA (DNA from the unborn baby) found in the mother’s bloodstream• Diagnosing inherited or spontaneous diseases, either as a result of symptoms, or because of family history (e.g. Duchenne muscular dystrophy)? Genetic fingerprinting PCR is used as a tool for genetic fingerprinting, also known as DNA profiling from a sample of blood or semen or from a hair root. Genetic fingerprints are profiles of specific stretches of DNA that vary from person to person. ? Detection and diagnosis of diseasesPCR-based diagnostics tests are accessible for identifying and/or evaluating various pathogens before standard serological laboratory tests (tests to detect the presence of antibodies) thus allowing early onset of the treatments. Examples of pathogens that can be identified:• HIV-1 (causes AIDS)• Hepatitis B and C infections (prompts liver malignancy)• Human Papillomavirus (cause cervical growth)• Chlamydia trachomatis (may prompt female infertility)• Neisseria gonorrhoeae (cause pelvic provocative illness in females)• Mycobacterium tuberculosis (causes tuberculosis)? Detection of infection in the environmentPCR is used to monitor and track the spread of infectious disease within an animal or human population. PCR can also be used to detect bacterial and viral DNA in the environment, for example looking at pathogens in water supplies.? Human Genome ProjectThe Human Genome Project (HGP) was an international scientific research project with the goal of determining the sequence of chemical base pairs which make up human DNA, and of identifying and mapping all of the genes of the human genome from both a physical and functional standpoint. The grouping of the billion bases in the HGP depended vigorously on PCR.Perhaps, PCR is the most influential of all techniques that has enabled gene cloning, sequencing of complex genomes, DNA fingerprinting and DNA-based diagnostics and many other techniques that were either inefficient or impossible before PCR. References:1.