PCR (Polymerase Chain Reaction)

 

PCR (Polymerase Chain Reaction) has become one of the most important tools in bioscience, diagnostics, and forensic science today. It is mainly used in two key areas: high-throughput PCR systems and microfluidic PCR devices for point-of-care (POC) applications. PCR, or Polymerase Chain Reaction, was invented in the 1980s by Kary Mullis, who later won the Nobel Prize in 1994 for his work. This technique has revolutionized biological research, making it easier to study basic biological processes and helping in areas like disease diagnosis and improving the genetics of plants and animals.

Over the past 100 years, few inventions have had as much impact as PCR, which has transformed biological and genetic research. The inventor, Kary Mullis, observed that while using the Sanger sequencing method to sequence a single-copy gene, the signals were weak because there wasn't enough DNA. By adding a step to separate the double-stranded DNA (dsDNA) into single strands (ssDNA) and introducing a reverse primer to define the length of the DNA to be copied, he was able to double the number of DNA copies. This process became the foundation of PCR, enabling DNA to be amplified in large quantities.

PCR was originally developed to detect mutations in the HBB gene, which causes sickle cell anemia. In this early project, scientists used radioactive-labeled DNA pieces and a method called restriction analysis to find inherited mutations. They chose sequences from the β-globin gene for the first PCR-based tests.

Advancements from PCR include:

  • Digital PCR
  • Droplet-based digital PCR (ddPCR)
  • Chip-based digital PCR (cdPCR)
  • Isothermal amplification

PCR is also crucial in techniques like Next-Generation Sequencing (NGS), and its role in molecular diagnostics is expected to grow significantly in the future.

Principles of polymerase chain reaction (PCR)

Recent advances in molecular methods have transformed how we detect and study microorganisms in various medical fields, like virology, microbiology, and dentistry. Among these methods, Polymerase Chain Reaction (PCR) has been a game-changer, helping to quickly identify pathogens, especially those that are hard to grow in the lab. In addition to regular PCR, Real-Time PCR has emerged as a major innovation, becoming increasingly important in both clinical diagnostics and research. Real-Time PCR is valued for its ability to provide both qualitative (yes/no) and quantitative (how much) results, making it a fast and reliable tool for detecting microorganisms.

PCR works by copying specific sections of DNA. It uses an enzyme called DNA polymerase, which plays a role in natural DNA replication within cells. In PCR, this enzyme creates a complementary DNA sequence by attaching to a small DNA fragment, called a primer, which marks the exact spot where copying should begin. The primers define the section of DNA to be replicated, leading to the creation of billions of copies of that particular sequence. This method has been extremely useful for gene analysis, diagnosing genetic diseases, and detecting pathogens like bacteria, viruses, and fungi. PCR can also be used for cloning DNA fragments to study gene activity, which has important uses in fields like forensic science.

Real time PCR

Real-Time PCR is a technique that accurately measures specific nucleic acids, making it very useful for diagnosing pathogens and genetic diseases. This method is highly sensitive and provides consistent results. It works by using a machine called a thermocycler, which has an optical system to detect fluorescent signals, and a computer with software to analyze the data. Different manufacturers offer machines with varying sample capacities, sensitivity levels, and methods for processing data. As the PCR process amplifies the DNA, the fluorescent dyes emit light in proportion to the amount of double-stranded DNA produced. The fluorescence is recorded during each cycle, showing how much PCR product is generated.

Applications of PCR

The discovery of Polymerase Chain Reaction (PCR) brought a major breakthrough in detecting microorganisms, making diagnoses more sensitive, precise, and accurate.

  • In virology, PCR helps detect and analyze viral DNA or RNA, giving us a deeper understanding of how viruses behave and cause infections.

  • In the fields of mycology and parasitology, PCR allows for early identification of fungi and parasites, which improves disease tracking and diagnosis—critical for identifying harmful pathogens.

  • In microbiology, PCR plays a key role in quickly detecting microorganisms like Mycobacterium tuberculosis, which is vital for timely and effective diagnostic tests.


Link to the Research Paper

Comments

Post a Comment

Popular posts from this blog

Application of Biotechnology in Cancer research

Image
  Cancer research and drug development have advanced significantly with the rise of new technologies like whole-genome sequencing, proteome analysis, and exome sequencing. These methods have provided fresh insights and information. In the early stages, drug discovery typically begins in academic settings, where researchers explore and lay the foundation for targeting specific proteins, pathways, or cellular networks to achieve therapeutic effects. Over the last decade, there has been significant progress in understanding cancer and developing targeted treatments. Molecular research has identified key mutations in various cancer types, allowing for the creation of drugs aimed at these specific targets. Another breakthrough was understanding that cells within the same tumor can vary in their characteristics, which explains why some treatments work differently for primary tumors compared to metastases in the same patient. Additionally, researchers have uncovered how tumor...
Read more

Foundations: How We Feel Hunger and Fullness 🧠

Image
  Our Brain’s Hunger Center: The Hypothalamus We rely heavily on a small but powerful brain region called the hypothalamus, which acts like a central appetite control panel. Within it, the ventromedial hypothalamus (VMH) contains two groups of neurons: One group tells us to eat more when energy is low, when hungry. Another tells us to stop eating when we’re full. Apart from taste, our insular cortex processes sensations of texture, mouthfeel, and even the pleasure of chewing. This is why crunching on celery feels more satisfying than sipping a smoothie—our brain uses that feedback to decide when to eat and when to stop eating. Key Hormones: Orchestrating Hunger and Satiety Ghrelin: The Hunger Hormone - Produced in the stomach, ghrelin rises when we haven’t eaten—giving that tell-tale “I need food” feeling. It’s tied to regular eating times, if we always eat lunch at noon, ghrelin shows up like clockwork. MSH & AgRP Neurons: Within the hypothalamus is the...
Read more

Steatotic liver disease - Causes & Treatment

Image
  Steatotic liver disease refers to disorders characterized by abnormal lipid accumulation in the liver (liver or hepatic steatosis). Steatotic liver disease includes what was previously known as non-alcoholic fatty liver disease (NAFLD), but is now referred to as metabolic dysfunction-associated steatotic liver disease. Steatotic liver disease also includes alcohol-related liver disease (ALD), MetALD, a new categorization for the overlap between MASLD and ALD, as well as unusual causes of liver steatosis. Steatotic liver disease affects around 30% of the global population and is mostly caused by obesity, type 2 diabetes, and alcohol consumption; however, only a small fraction of those with steatotic liver disease develop cirrhosis. The presence and advancement of liver fibrosis caused by hepatic inflammation is the most reliable predictor of liver death across the whole range of steatotic liver disorders. A combination of recent advances in widely available biomarkers ...
Read more