International Journal of Multidisciplinary Research and Publications ISSN (Online): 2581-6187 191 Amna Arooj and Hifz Ur Rehman, “Evolution and Future of DNA Extraction Procedure,” International Journal of Multidisciplinary Research and Publications (IJMRAP), Volume 7, Issue 5, pp. 191-197, 2024. Evolution and Future of DNA Extraction Procedure Amna Arooj 1 , Hifz Ur Rehman 2 , Hafiz Muhammad Abbas Malik 3 , Muhammad Shahid Cholistani 2 , Sabahat Shakoor 2 , Irshad Ahmed 2 , Areesha Murtaza 1 , Wishma Ashfaq 1 , Muhammad Hashim Raza 2 , Samiullah Malik 4* 1 The Department of Forensic Science, Faculty of Allied Health Sciences, The Islamia University of Bahawalpur, Bahawalpur 63100, Punjab, Pakistan 2 Institute of Biological Sciences, Khwaja Fareed University of Engineering and Information Technology, Rahim Yar Khan 64200, Punjab, Pakistan 3 The Department of Pharmaceuticals, Faculty of Pharmacy, Islamia University of Bahawalpur, Bahawalpur 63100, Punjab, Pakistan. 4 Sir Sadiq Muhammad Khan Abbasi Post Graduate Medical College, Faculty of Medicine & Allied Health Sciences, The Islamia University of Bahawalpur, Bahawalpur 63100, Punjab, Pakistan *Corresponding Author: Samiullah Malik, E-mail: samiullah.mushtaq@iub.edu.pk Abstract—DNA extraction holds paramount importance in forensic science, genetic research, and clinical diagnostics. It makes the DNA used for analysis in different applications including criminal investigations. After the mid-20 th century, DNA extraction techniques evolved with early methods such as phenol-chloroform extraction which were labor-intensive and used hazardous chemicals. Over time newer techniques like silica-based, magnetic bead-based, and automated systems emerged making DNA extraction more efficient and accessible. Recent innovations include microfluidic and nanotechnology-based methods which aim to enhance yield and sensitivity, particularly for challenging samples like touch DNA. Still, limitations persist especially in handling low-template or degraded DNA samples. Current methods may lack the sensitivity or purity required for reliable analysis of trace DNA posing challenges in extracting usable DNA from minimal or compromised samples. Future innovations may yield breakthroughs that enable reliable DNA extraction even from a minute quantity of samples. Keywords— Automated systems, chelex extraction, DNA extraction, magnetic-beads, microfluidic, nanotechnology, Phenol-Chloroform, Salting-Out, silica-column, touch DNA. I. INTRODUCTION DNA extraction has served as the foundational step in molecular biology for over half a century facilitating a range of scientific fields including genetics, clinical diagnostics, and forensic science. The fundamental goal of DNA extraction is to isolate pure DNA from cellular material in a manner that preserves the quality and quantity of the extracted nucleic acid. The method chosen often depends on the nature and quantity of the sample as well as the intended downstream application which may require high sensitivity, minimal contamination, or the preservation of DNA integrity. [1][2] Early methods like the phenol-chloroform extraction developed in the 1950s were based on organic solvents that separate nucleic acids from proteins. While effective these techniques involved hazardous chemicals and multiple steps that posed challenges for scalability and routine use. Over time techniques evolved with the development of salting-out methods in the 1980s which provided a safer and faster alternative but were less effective for certain types of samples [2]. In the 1990s silica-based and magnetic bead-based methods revolutionized the extraction landscape by providing more user- friendly high-throughput alternatives that increased purity and yield. These methods were particularly useful for forensic applications as they allowed the extraction of DNA from degraded or trace biological samples [3]. By the 2000s automated extraction systems became widely available enabling consistent and high-throughput DNA isolation with minimal human intervention ideal for both clinical and forensic settings [4]. Recent advancements in the 2010s and 2020s have focused on microfluidics and nanotechnology to improve extraction efficiency and sensitivity further. These methods hold promise for ultra-low quantity samples such as touch DNA—DNA left by incidental contact which typically exists in trace amounts and is prone to environmental degradation [5]. In forensic science, DNA extraction plays a critical role in identifying suspects, linking individuals to crime scenes, and exonerating the innocent. Touch DNA, however, presents unique challenges due to the small amounts of DNA present and the risk of contamination. Effective extraction techniques for touch DNA are essential for obtaining reliable forensic results yet current methods still face limitations in yield and sensitivity. II. DNA EXTRACTION DNA extraction is the process of isolating DNA from cells or tissues. It’s a critical first step in various biological and forensic applications as it allows scientists to study genetic material in a purified form. The extraction process involves breaking down the cell membrane, separating DNA from other cellular components, and purifying it for analysis. Once extracted, DNA can be used for tasks like genetic testing, forensic identification, research on genetic diseases, and analyzing biological relationships. [6] Typical DNA extraction involves (a) cell lysis, which is breaking open cells to release their DNA. This can be done chemically using detergents or enzymes or physically by grinding or blending. (b) Removal of Proteins and Other Contaminants: Proteinase K or other enzymes help digest proteins while additional reagents like salt solutions help