ChemTalk

Frederick Sanger: The Man Behind DNA Sequencing

Frederick Sanger, the inventor of Sanger Sequencing
Frederick Sanger

The central dogma of biology is what allows life to march on. It is a chain of production, in which DNA is transcribed into RNA, which is translated into protein. This means that each gene’s specific sequence of bases dictates the structure and function of its associated protein. Proteins carry out processes that keep organisms alive. Today, finding those exact sequences and analyzing them is an important part of biology research called DNA sequencing. Frederick Sanger, the inventor of Sanger Sequencing, was a key figure in establishing the central dogma. He is considered the father of DNA sequencing. Today, labs across the world have their samples sequenced to check the results of their genetic edits, ascertain the guilt of a suspect, or pin down an organism’s place on an evolutionary tree.

Discovering the Amino Acid Sequence of Insulin

Sanger began his research career at Cambridge University in the 1940s. He was investigating the amino acid composition of insulin, one of the only proteins readily available in a pure form at the time. Sanger discovered the amino acid sequence of two forms of bovine insulin. While doing so, he realized that the sequences needed to be precise and consistent. The two chains were held together by disulfide bonds between Cysteine amino acids. If the peptide sequence changed at all, the chains would not stay together and the protein would not function. Before this landmark discovery, most researchers thought that proteins were amorphous and inconsistent. Sanger’s finding laid the foundation for the central dogma of molecular biology. The enormous breakthrough earned Sanger his first Nobel prize.

This graphic shows the disulfide bonds (in red) that hold the A and B chains of insulin together. Disulfide bonds can only occur between cysteine residues, so it is crucial that the sequence of both chains stays consistent.
Another visualization of the interlinked chains of insulin

Sequencing 5S RNA

Sanger then turned toward the second stop in the developing central dogma, RNA. While his lab was not the first to successfully sequence RNA, they accidentally made another discovery that would be foundational to the young field of genetics. They sequenced RNA from E. coli called 5S ribosomal RNA. Ribosomal RNA, or rRNA, makes up part of the ribosome. The ribosome is an important organelle that serves as the site of translation from RNA to protein. Today, that rRNA sequence is useful in classifying and identifying bacteria.

Sanger Sequencing

Finally, in 1977, Sanger introduced dideoxy chain-termination DNA sequencing. The method involves copying single-stranded DNA, but adding dideoxynucleotides (ddNTPs) to the mix. ddNTPs cause early termination of DNA replication. Researchers run four different reactions, one with each type of dideoxynucleotide (A, C, T, and G) added. The procedure results in several single strands of DNA, each varying in length by a single nucleotide. The last nucleotide in the chain will be a radio-labeled ddNTP. 

Researchers then sort the strands from all four reactions by size using a polyacrylamide gel, with each lane receiving either the A, C, T, or G ddNTP treatment. The gel allows shorter strands to travel faster and farther than long strands. This means longer strands will produce dark bands toward the top of the gel, and smaller strands produce bands toward the bottom. Thus, scientists can easily ascertain the sequence of the strand by looking at the bands and assessing whether the labeled nucleotide is A, C, T,  or G.

A polyacrylamide gel showing the results of Sanger sequencing
This image shows a gel obtained after using Sanger sequencing. Each band represents a DNA strand of a different length that ends with a certain ddNTP. We can see that the sequence begins, from the top of the gel, “TCGTTATCTTCC…”

This method, now known as the Sanger method, was the predominant DNA sequencing method. Sanger earned his second Nobel prize for the innovation of DNA sequencing in 1980. Sanger’s method is remembered as a breakthrough in biology, even though newer and more efficient sequencing methods have overtaken it. Sanger sequencing was part of a crucial foundation for the future of genomics and countless other fields of science.