The Usage of Sanger sequencer
November 07, 2023
The Sanger sequencing method, also known as chain termination sequencing, is a widely used technique in molecular biology and genetics for determining the order of nucleotide bases in a DNA molecule. It was developed by Frederick Sanger and his colleagues in the 1970s and earned Sanger the Nobel Prize in Chemistry in 1980.
The Sanger sequencing Genetic Analyzer method relies on the incorporation of chain-terminating dideoxynucleotides (ddNTPs) during DNA synthesis. These ddNTPs lack the 3'-OH group necessary for further DNA chain elongation, effectively terminating the DNA strand at specific positions. The DNA is first divided into multiple identical fragments, and each fragment is subjected to DNA synthesis with normal nucleotides (dNTPs) and a small amount of one of the four ddNTPs labeled with different fluorophores. As a result, the DNA synthesis reaction produces a set of fragments of varying lengths, each terminating with a specific ddNTP. The fragments are then separated by capillary electrophoresis, allowing for the determination of the DNA sequence based on the fluorescent signals emitted by the different labeled ddNTPs.
DNA Sequencer Display
The Sanger Sequecing Instrument, also known as a DNA sequencer or genetic sequencer, is the instrument used for automated Sanger sequencing. It consists of several key components, including a thermal cycler, a capillary electrophoresis system, a fluorescence detection system, and a computer for data analysis and interpretation. The thermal cycler is used to carry out the DNA synthesis reaction and control the temperature cycles required for enzymatic reactions. The capillary electrophoresis system separates the DNA fragments based on their size and charge and detects the fluorescent signals emitted by the labeled ddNTPs. This information is then processed by the fluorescence detection system, which converts the signals into a digital format. The computer software analyzes the data and generates the DNA sequence output.
The Sanger sequencer has played a crucial role in various fields of biological research, including genome sequencing, genetic analysis, mutation detection, and DNA cloning. With its high accuracy, relatively low cost, and scalability, the Sanger sequencing method has become a fundamental tool in molecular biology laboratories worldwide.
