Sanger sequencing, also known as dideoxy sequencing, is a method for determining the sequence of nucleotides in a DNA molecule. The basic principle of Sanger sequencing is to use DNA polymerase to synthesize DNA chains in the presence of dideoxy nucleotides, which are modified nucleotides that lack a 3'-hydroxyl group, which is required for the formation of a phosphodiester bond between nucleotides. When a dideoxy nucleotide is incorporated into a growing DNA chain, it terminates the chain and prevents further extension.
In Sanger sequencing, multiple DNA fragments are synthesized in parallel, each starting from a different position along the target DNA strand and using a different dideoxy nucleotide. The resulting DNA chains of different lengths are separated by gel electrophoresis, and the sequence of nucleotides can be determined by comparing the patterns of bands on the gel.
Next generation sequencing (NGS) technologies, such as Illumina, PacBio, and Oxford Nanopore sequencing, have revolutionized DNA sequencing by allowing the simultaneous sequencing of millions or billions of DNA fragments in a single run. NGS technologies are faster and more cost-effective than Sanger sequencing, but have lower accuracy and read lengths compared to Sanger sequencing.
Which method is most preferred depends on the specific requirements of the research project. Sanger sequencing is still commonly used for a variety of applications, particularly when high accuracy and long read lengths are required. For large-scale genome projects, NGS technologies are more practical and cost-effective. However, for specific applications such as validation of genomic variants, Sanger sequencing may still be preferred due to its high accuracy and specificity.