You decide to determine the complete nucleotide sequence of

You decide to determine the complete nucleotide sequence of the gene by DNA sequencing using fluorescent nucleotides. Which of the below nucleotides(s) (1/2/3) are used in DNA sequencing reaction and in what type of sequencing? Show all that apply. 2 and 3 are used in DNA sequencing reaction.Which of the below nucleotides(s) (1/2/3) would you fluorescently tag for DNA sequencing and why

Solution

Answer:

For sequencing DNA dideoxynucleotide (ddNTP) is used (figure 3). Dideoxynucleotides (ddATP, ddTTP, ddGTP, ddCTP) are labeled with different color of the dye for sequencing. This method is called as Sanger sequencing.

Since dideoxynucleotide lacks a hydroxyl group on the 3\'-carbon therefore, once the polymerase incorporates a ddNTP to the chain, the chain terminates as Polymerase is unable to add further nucleotides to extend the chain.