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What is the Sanger method?

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The Sanger method is a technique for sequencing DNA that uses dideoxynucleotides to make short DNA sequences of different lengths that can be read out in order using gel electrophoresis. Dideoxynucleotides are similar to the regular deoxynucleotides (the A, G, C and T in DNA) except they are missing the -OH group that forms bonds with the next nucleotide. This omission prematurely ends the DNA strand.

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The Sanger method requires the DNA to be replicated in four different tubes. Each tube contains a polymerase to carry out the DNA replication, the template DNA for sequencing, all four of the regular deoxynucleotides (dNTP) and some dideoxynucleotide (ddNTP) from one of the bases. This means each of the four tubes has A, G, C and T as dNTPs and one of the bases as a ddNTP.

Replication is allowed to continue for a period of time, and then the strands of DNA are run on a gel via electrophoresis. There is a chance that every time one of the nucleotides that is identical to the dideoxynucleotides is incorporated into the DNA, the ddNTP is incorporated instead, ending the DNA strand prematurely. Each tube then contains pieces of DNA that end with ddNTPs, and separating these strands by size makes it possible to see where A, G, C or T appears in the sequence.

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