Biochemistry Questions Biochemistry Questions / Explain the principle of Sanger dideoxy sequencing and its significance in the Human Genome Project.

Sanger dideoxy sequencing was, and still is, the dominant sequencing technology. The Human Genome Project (HGP) used this technology, combined with fluorescent terminator (dNTPs), to take advantage of its long read lengths. This allowed the HGP to overcome short repeat sequences found in the human genome, enabling a longer (but still discontinuous) sequence to be produced. In dideoxy sequencing, chain terminating dNTPs are used to produce DNA strands of varying lengths. Although this technology originally used radiolabelled dNTPs and four reactions (one of each A, T, C, and G), the technology had matured to using fluorescent dNTPs and a single reaction. By using capillary gel electrophoresis, the fluorescence of the DNA strands can be recorded, in the order of shortest to longest. This allowed for the long 800bp reads used in the HGP. Without high-throughput sequencing techniques, the HPG would not have been able to be carried out. The large scale use of sequencing here resulted in major advancements, including next generation sequencing. This allows for the simultaneous reading of many thousands of DNA strands, enabling for faster Sanger sequencing as well as reducing costs.