Editing Sequencing (section)

===Pyrosequencing===
{{main|Pyrosequencing}}
The pyrosequencing method is based on the detection of the pyrophosphate release on nucleotide incorporation. Before performing pyrosequencing, the DNA strand to sequence has to be amplified by PCR. Then the order in which the nucleotides have to be added in the sequencer is chosen (i.e. G-A-T-C). When a specific nucleotide is added, if the DNA polymerase incorporates it in the growing chain, the pyrophosphate is released and converted into ATP by ATP sulfurylase. ATP powers the oxidation of luciferase through the luciferase; this reaction generates a light signal recorded as a pyrogram peak. In this way, the nucleotide incorporation is correlated to a signal. The light signal is proportional to the amount of nucleotides incorporated during the synthesis of the DNA strand (i.e. two nucleotides incorporated correspond to two pyrogram peaks). When the added nucleotides aren't incorporated in the DNA molecule, no signal is recorded; the enzyme apyrase removes any unincorporated nucleotide remaining in the reaction.
This method requires neither fluorescently-labelled nucleotides nor gel electrophoresis.
Pyrosequencing, which was developed by Pål Nyrén and Mostafa Ronaghi DNA, has been commercialized by Biotage (for low-throughput sequencing) and 454 Life Sciences (for high-throughput sequencing). The latter platform sequences roughly 100 [[megabase]]s [now up to 400 megabases] in a seven-hour run with a single machine.  In the array-based method (commercialized by 454 Life Sciences), single-stranded DNA is annealed to beads and amplified via [[454 Life Sciences#DNA library preparation and emPCR|EmPCR]].  These DNA-bound beads are then placed into wells on a fiber-optic chip along with [[enzymes]] which produce light in the presence of [[adenosine triphosphate|ATP]].  When free nucleotides are washed over this chip, light is produced as ATP is generated when nucleotides join with their complementary [[base pair]]s.  Addition of one (or more) nucleotide(s) results in a reaction that generates a light signal that is recorded by the CCD camera in the instrument. The signal strength is proportional to the number of nucleotides, for example, homopolymer stretches, incorporated in a single nucleotide flow.  [https://web.archive.org/web/20080318163514/http://www.454.com/]