Next-generation sequencing: Advantages, disadvantages, and future

Abstract

It has been more than 35 years since the development of the groundbreaking method for DNA sequencing by Frederick Sanger and colleagues. This revolutionary study triggered the improvement of new methods that have provided great opportunities for low-cost and fast DNA sequencing. Strikingly after the Human Genome Project, the time interval between each sequencing technology started decreasing while amount of scientific knowledge has continued growing exponentially. Considering Sanger sequencing as the first generation, new generations of DNA sequencing have been introduced consequently. The development of the next-generation sequencing (NGS) technologies has contributed to this trend substantially by reducing costs and producing massive sequencing data. Hitherto, four sequencing generations have been defined. Second-generation sequencing that is currently the most commonly used NGS technology consists of library preparation, amplification, and sequencing steps while in third-generation sequencing, individual nucleic acids are sequenced directly in order to avoid biases and have higher throughput. Recently described fourth-generation sequencing aims conducting genomic analysis directly in the cell. Classified to different generations, NGS has led to overcome the limitations of conventional DNA sequencing methods and has found usage in a wide range of molecular biology applications. On the other hand, plenty of technical challenges, which need to be deeply analyzed and solved, emerged with these technologies. Every sequencing generation and platform, by reason of its methodological approach, carries characteristic advantages and disadvantages which determine the fitness for certain applications. Thus, assessment of these features, limitations, and potential applications help shaping the studies that will determine the route of omic technologies.