Introduction
DNA sequencing is the process of determining the exact order of nucleotides (A, T, C, G) in a DNA molecule. It plays a critical role in understanding genetics, diagnosing diseases, and advancing biotechnology. In this answer, we will discuss the principles and methods of DNA sequencing and explain its importance in genetic studies.
Principles of DNA Sequencing
The basic principle of DNA sequencing is to identify the order of bases in a DNA strand. Sequencing involves copying DNA in a way that allows scientists to detect each base added during the process. Different sequencing methods use various technologies to achieve this.
Traditional Method: Sanger Sequencing
Sanger sequencing, also called the chain termination method, was developed by Frederick Sanger in 1977 and was the first widely used sequencing technique.
Steps in Sanger Sequencing:
- DNA is copied using DNA polymerase, regular nucleotides (dNTPs), and special chain-terminating nucleotides (ddNTPs).
- When a ddNTP is added, the DNA strand stops growing.
- Fragments of varying lengths are created and separated using gel or capillary electrophoresis.
- Each ddNTP is labeled with a fluorescent dye, allowing the sequence to be read by a detector.
Advantages: Accurate and reliable; suitable for sequencing small DNA fragments.
Limitations: Time-consuming and expensive for large-scale sequencing.
Modern Methods: Next-Generation Sequencing (NGS)
Next-generation sequencing allows millions of DNA fragments to be sequenced simultaneously.
1. Illumina (Sequencing by Synthesis)
- DNA is broken into short fragments and attached to a flow cell.
- DNA is amplified, and bases are added one at a time.
- Each added base emits a fluorescent signal that is recorded by a camera.
2. Ion Torrent
- DNA polymerase adds nucleotides, and a hydrogen ion is released with each base added.
- The change in pH is measured to identify the base.
3. PacBio and Oxford Nanopore (Long-read Sequencing)
- Read long DNA sequences in real-time.
- Useful for complex genomes and structural variation detection.
Why is DNA Sequencing Important in Genetic Studies?
1. Understanding Genes and Mutations
Sequencing helps identify genes and mutations that cause genetic diseases. It is used to study inherited disorders and cancer.
2. Personalised Medicine
By sequencing an individual’s genome, doctors can customize treatment based on their genetic profile.
3. Evolution and Ancestry
Comparing DNA sequences across species helps trace evolutionary history and study human ancestry.
4. Identifying Pathogens
Sequencing helps detect and identify bacteria and viruses in infections, especially in pandemics like COVID-19.
5. Agricultural Biotechnology
Sequencing plant and animal genomes helps improve traits like disease resistance and yield in crops and livestock.
6. Functional Genomics
Researchers study how genes work and interact by sequencing and analyzing DNA regions.
Conclusion
DNA sequencing is a powerful tool that helps unlock the secrets of life. From the traditional Sanger method to modern next-generation sequencing, the technology has evolved to offer faster, cheaper, and more accurate results. It is essential in genetic studies for understanding health, disease, evolution, and more. As sequencing becomes more accessible, its applications in medicine, agriculture, and research will continue to grow.