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Microfluidic Sanger sequencing is a lab-on-a-chip application for DNA sequencing, in which the Sanger sequencing steps (thermal cycling, sample purification, and capillary electrophoresis) are integrated on a wafer-scale chip using nanoliter-scale sample volumes. This technology generates long and accurate sequence reads, while obviating many ...
For Sanger sequencing, either cloning procedures or PCR are required prior to sequencing. In the case of next-generation sequencing methods, library preparation is required before processing. [ 162 ] Assessing the quality and quantity of nucleic acids both after extraction and after library preparation identifies degraded, fragmented, and low ...
DNA sequencing is the process of determining the nucleotide order of a given DNA fragment. So far, most DNA sequencing has been performed using the chain termination method developed by Frederick Sanger. This technique uses sequence-specific termination of a DNA synthesis reaction using modified nucleotide substrates.
The classical shotgun sequencing was based on the Sanger sequencing method: this was the most advanced technique for sequencing genomes from about 1995–2005. The shotgun strategy is still applied today, however using other sequencing technologies, such as short-read sequencing and long-read sequencing.
The AB370A was able to sequence 96 samples simultaneously, 500 kilobases per day, and reaching read lengths up to 600 bases. This was the beginning of the "first generation" of DNA sequencers, [2] [3] which implemented Sanger sequencing, fluorescent dideoxy nucleotides and polyacrylamide gel sandwiched between glass plates - slab gels. The next ...
The sequencing occurs for millions of clusters at once, and each cluster has ~1,000 identical copies of a DNA insert. [12] The sequence data is analyzed by finding fragments with overlapping areas, called contigs, and lining them up. If a reference sequence is known, the contigs are then compared to it for variant identification.
Although Maxam and Gilbert published their chemical sequencing method two years after Frederick Sanger and Alan Coulson published their work on plus-minus sequencing, [2] [3] Maxam–Gilbert sequencing rapidly became more popular, since purified DNA could be used directly, while the initial Sanger method required that each read start be cloned for production of single-stranded DNA.
In 1991 Adams and co-workers coined the term expressed sequence tag (EST) and initiated more systematic sequencing of cDNAs as a project (starting with 600 brain cDNAs). [8] The identification of ESTs proceeded rapidly, millions of ESTs now available in public databases (e.g. GenBank).
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