Sanger Sequencing

Definition

Sanger sequencing, also known as chain termination sequencing, is a method used to determine the nucleotide sequence of DNA. This technique revolutionized genomics by allowing for accurate and efficient sequencing of DNA, making it essential in bioinformatics and genomics workflows for understanding genetic information.

5 Must Know Facts For Your Next Test

1. Sanger sequencing was developed by Frederick Sanger in 1977 and was the first method used to sequence DNA accurately.
2. The process relies on the use of dideoxynucleotides, which terminate DNA strand elongation, allowing the generation of fragments of varying lengths that can be separated by size.
3. This technique is widely used in various applications, including genetic testing, cloning, and sequencing of PCR products.
4. Sanger sequencing remains a gold standard for validating sequences obtained from next-generation sequencing technologies due to its high accuracy.
5. The method can sequence relatively short stretches of DNA, typically up to about 1,000 base pairs in length, making it ideal for specific target regions.

Review Questions

• How does Sanger sequencing differ from other DNA sequencing methods in terms of technology and application?
  • Sanger sequencing differs from other methods like Next-Generation Sequencing (NGS) primarily in its approach and scale. While Sanger uses chain-terminating dideoxynucleotides to create fragments for size separation, NGS employs parallel processing to sequence millions of fragments simultaneously. Sanger is often used for smaller, targeted sequences or for validation of NGS results due to its high accuracy, while NGS is preferred for whole-genome sequencing or large-scale studies.
• Discuss the role of dideoxynucleotides in Sanger sequencing and how they facilitate the determination of DNA sequences.
  • Dideoxynucleotides (ddNTPs) play a critical role in Sanger sequencing by causing premature termination of DNA strand synthesis. When incorporated into a growing DNA strand during replication, ddNTPs lack a hydroxyl group at the 3' position, preventing further nucleotide addition. This results in fragments of different lengths that correspond to the sequence of the original template DNA. By analyzing these fragments through capillary electrophoresis or gel electrophoresis, researchers can determine the specific order of nucleotides in the DNA sequence.
• Evaluate the impact of Sanger sequencing on the field of genomics and bioinformatics workflows since its inception.
  • Since its introduction, Sanger sequencing has profoundly impacted genomics and bioinformatics by providing a reliable method for accurately determining DNA sequences. Its ability to generate high-quality data laid the groundwork for subsequent advancements in genomic research and paved the way for large-scale projects like the Human Genome Project. Additionally, Sanger sequencing's high accuracy makes it essential for validating results obtained from newer technologies like Next-Generation Sequencing (NGS), ensuring data integrity in bioinformatics workflows. This interplay between Sanger and NGS exemplifies how foundational techniques continue to influence modern scientific practices.

Related terms

DNA Polymerase: An enzyme that synthesizes new DNA strands by adding nucleotides to a growing chain during DNA replication and sequencing.

Dideoxynucleotides (ddNTPs): Modified nucleotides used in Sanger sequencing that cause termination of the DNA strand during replication, enabling the determination of DNA sequences.

Next-Generation Sequencing (NGS): A group of advanced sequencing technologies that allow for the rapid sequencing of large amounts of DNA, often used in conjunction with Sanger sequencing for more comprehensive analyses.