Describe the Sanger method of DNA sequencing

Question

What will be an ideal response

Answer 1

Complete DNA replication will occur when normal deoxynucleotides (dNTP) are used. When dideoxynucleotides
are incorporated into the growing chain (ddNTP), the chain terminates replication because the hydroxyl group is
missing that is necessary for the next nucleotide to be added. If DNA replication occurs in a mixture composed
mostly of normal dNTPs but also a small amount of ddNTPs, then DNA chains of different lengths will be
produced. If one wishes to sequence a particular piece of DNA, then it is aliquoted into four parts. A primer is
added to each aliquot because DNA polymerase needs a primer to start replication, and this insures that DNA
replication begins at the same point in each piece of DNA. In one part, replication occurs with normal dNTPs plus a
small amount of dideoxyadenine (ddATP); in the next, dNTPs plus dideoxythymine (ddTTP) followed by
dideoxyguanine (ddGTP) and dideoxycytosine (ddCTP). When the DNA fragments are analyzed from each aliquot,
the point at which any particular chain terminated corresponds to the dideoxynucleotide added to that aliquot.
When the fragments are analyzed together using high resolution denaturing polyacrylamide gel electrophoresis,
the sequence of the original fragment can be determined.