Written by Sanger sequencing determines the nucleotide bases’ sequence in a DNA piece. It is also called the chain termination method and was developed by Frederick Sanger in 1977. Sanger sequencing has an accuracy of up to 99.99%; therefore, it is considered the most effective method of determining DNA sequences, including those sequenced by NGS (next-generation sequencing). Sanger sequencing can determine the sequences of small DNA fragments up to 1000 bp in length. These fragments are the ones used to connect the bigger DNA fragments and, ultimately, the entire chromosomes.
Sanger Sequencing Versus NGS
Both are molecular biology lab services, although the establishment of NGS technologies has revolutionized genomics research. NGS can sequence whole genomes and over 1oo genes with moderate-input DNA. However, Sanger sequencing is widely used due to several merits, including its accuracy, therefore suitable for validation sequencing and cost-effectiveness for single genes sequencing.
How Sanger Sequencing Works
This procedure is categorized among microbiology tests. The DNA to be sequenced, DNA primer to match with the template DNA starts the process of DNA synthesizing. It is usually done in the existence of four deoxy nucleotide triphosphates (dNTPs): A, C, G and T, whereby the polymerase stretches the primer by addition of the matching dNTP to the template DNA length. The four dNTPs: dATP, dTTP, dGTP, and dTTP, terminate the synthesis reaction and determine the nucleotide incorporated into the series of nucleotides.
The dNTPs have an oxygen element detached from the ribonucleotide and thus cannot link with the next nucleotide. The sequence of DNA is determined when the synthesis produces a reaction loading the products into four lanes of one gel, with the action of the assorted chain-terminating nucleotide and gel electrophoresis which separates the denatured fragments.
