Question

In Sanger sequencing after the strands are separated a strand is copied in a reaction mix containing small amounts of a ______________ nucleotide which causes synthesis to stop at occurrences of that nucleotide.

Answer 1

In Sanger sequencing, the incorporation of a dideoxynucleotide into a growing DNA strand causes chain termination, resulting in DNA fragments of various lengths, which are crucial for determining DNA sequences.

Step-by-step explanation:

In Sanger sequencing, after the DNA strands are separated, one strand is copied in a reaction mix containing small amounts of a dideoxynucleotide (ddNTP). This particular nucleotide causes DNA synthesis to stop at occurrences of that nucleotide. The ddNTPs are critical for Sanger sequencing; they lack the 3'-hydroxyl group (-OH) necessary for the formation of a phosphodiester bond with the next nucleotide, thus terminating DNA chain elongation whenever they are incorporated into the growing DNA strand. Four different ddNTPs are used, each corresponding to one of the four normal bases of DNA: adenine (ddATP), thymine (ddTTP), cytosine (ddCTP), and guanine (ddGTP), with each type added to separate reaction tubes. This process creates a variety of DNA fragments of different lengths, which can be separated via gel electrophoresis and subsequently sequenced.