In the realm of molecular biology, the accurate detection and quantification of nucleic acids has always been a challenge. In this context, to address the limitations of traditional polymerase chain reaction (PCR) methods, real time polymerase chain reaction systems emerge as a solution. This revolutionary tool offers unprecedented accuracy, sensitivity, and efficiency. In this blog, we will explore the developments in this field and delve into the exciting potential they hold.

What are Real Time Polymerase Chain Reaction Systems?

Real-time polymerase chain reaction, or real-time PCR, is a laboratory technique used in molecular biology. Real-time PCR is also referred to as quantitative PCR (qPCR) or real-time quantitative PCR. It is a variation of the traditional PCR method that allows for the monitoring of DNA amplification in real time rather than at the end of the reaction. This technique is particularly useful for investigating gene expression and quantifying specific nucleic acid sequences.

Real-time PCR has revolutionized molecular diagnostics and has numerous applications in fields such as food safety, public health, and the detection of pathogens like the SARS-CoV-2 virus. According to Kings Research, the real time polymerase chain reaction systems market is expected to reach a valuation of $7.01 billion by 2030.

What are the 4 Steps of Real-Time Polymerase Chain Reaction?

Real-time PCR involves four main steps to gain accurate quantification of DNA or RNA, including:

1. Reverse Transcription (RT)

The first step in RT-PCR is transforming RNA to complementary DNA (cDNA) using reverse transcription. This step is necessary because real-time PCR primarily detects and amplifies DNA rather than RNA. It uses reverse transcriptase enzymes, which synthesize cDNA from the RNA template.

2. Primer Design and Amplification

After reverse transcription, the next step involves setting up amplification reactions with PCR reagents and unique or custom primers. Primers are short DNA sequences that bind to specific regions of the target DNA, or cDNA. They define the region to be amplified during the PCR process. The amplification step uses a DNA polymerase enzyme to replicate the target DNA or cDNA, resulting in the exponential amplification of the specific genes of interest.

3. Fluorescent Detection

Real time polymerase chain reaction systems utilize fluorescent reporters to detect and quantify the amplified DNA or cDNA in real time. Two commonly used fluorescent reporters are SYBR Green and TaqMan probes. They both emit fluorescence when they bind to the target DNA or cDNA during the amplification process.

4. Data Analysis

The final step involves running the amplification reactions on real-time PCR instruments and analyzing the collected data using proprietary software. The software measures the fluorescence emitted during each amplification cycle and calculates the cycle threshold (Ct) value to determine the initial concentration of the target DNA or cDNA in the sample.

Real-life Applications of Real Time Polymerase Chain Reaction Systems

Here are some real-life applications of the qPCR system:

Medical Diagnostics: qPCR is widely used in medical diagnostics to detect and diagnose various diseases. It can identify the presence of pathogens, such as bacteria or viruses, in patient samples. PCR-based tests are commonly used for diagnosing infectious diseases, like COVID-19.
Genetic Testing: PCR is an essential tool in genetic testing. It can be used to identify genetic mutations associated with inherited diseases or to determine an individual's genetic predisposition to certain conditions.
DNA Cloning: PCR is routinely used in DNA cloning, a process that involves making copies of a specific DNA sequence. PCR allows researchers to amplify the desired DNA fragment, which can then be inserted into a vector for further manipulation and study.

Role of Real Time Polymerase Chain Reaction Systems in Forensic Analysis

RT-PCR system is a powerful and versatile tool that has revolutionized molecular biology and diagnostics. Its ability to detect and quantify genetic material in real time has made it an invaluable tool for a wide range of applications, including gene expression analysis, pathogen detection, and genotyping.

With its high sensitivity, specificity, and speed, RT-PCR has greatly contributed to our understanding of biological processes and has become an essential tool in both research and clinical settings. One of the best examples is the QuantStudio Real-Time PCR System by Thermo Fisher Scientific.

The systems detect gene expression changes as low as 1.5-fold, supporting various genomic applications. It has a user-friendly touchscreen, intuitive software, and a remote setup that helps monitor and share, reduce contamination, and increase productivity.

Anitoa's New Real-Time Polymerase Chain Reaction Modular System

Anitoa Systems recently launched the MAx16, an automation-ready real-time qPCR modular system. This fully integrated four-plex module with 16-well capacity is suitable for various applications, including sample or library preparation for DNA sequencing and integrated sample extraction to amplify and detect nucleic acid test systems.

The system has a fast thermal cycler that runs a complete RT-PCR protocol in less than 30 minutes. The system uses an electrically actuated heat lid for automation and gives a digital interface to the main automation system controller. The system has the potential to be integrated into a compact desktop robotic fluid-handling automation system easily.

Is AI Surpassing Ream-Time Polymerase Chain Reaction Test?

Australian researchers have developed an AI system that can detect COVID-19 from chest X-rays with over 98% accuracy, potentially surpassing the current RT-PCR test. The system uses a deep learning-based algorithm called Custom Convolutional Neural Network (Custom-CNN) to quickly distinguish between COVID-19 cases, normal cases, and pneumonia in X-ray images.

The system could be particularly beneficial in countries with high COVID-19 levels and shortages of radiologists. The system eliminates the need for manual search for biomarkers, allowing for faster and more accurate diagnosis. The system could prove beneficial in cases where PCR tests or rapid antigen tests show negative results.

In a Nutshell

Real time polymerase chain reaction systems are an innovative and robust instrument that has shifted molecular biology and diagnostics. Its ability to detect and quantify genetic material in real time has made it an invaluable tool for a wide range of applications, including gene expression analysis, pathogen detection, and genotyping.

With advancements, AI and RT-PCR have greatly contributed to our understanding of biological processes and have become an essential tool in both research and clinical settings.

Understanding the Role of Real Time Polymerase Chain Reaction Systems