B) Dideoxyribose blocks elongation
Growing fe DNA chain
Ribose 2'-deoxyribose 2',3'-dideoxyribose
FIGURE 24.05 Dideoxyribose, Deoxyribose and Ribose ddR
No 3' reactive hydroxyl group; elongation blocked
(A) The structures of ribose, deoxyribose, and dideoxyribose differ in the number and location of hydroxyl groups on the 2' and 3' carbons. (B) DNA poly merase cannot add another nucleotide to a chain ending in dideoxyribose because its 3' carbon does not have a hydroxyl group.
A) Random termination at "G" positions
Original sequence: TCGGACCGCTGGTAGCA
Mixture of chains terminated at G using mixtures of GTP and ddGTP:
4. TCiGACC CTG
6. TCiGACC CT ,G TAG
FIGURE 24.06 Chain Termination by Dideoxynucleotides
(A) During the sequencing reaction, DNA polymerase makes multiple copies of the original sequence. Sequencing reaction mixtures contain artificial dideoxynucleotides that terminate growing DNA chains. The example here shows the G reaction, which includes triphosphates of both deoxyguanosine (dG) and dideoxyguanosine (ddG). Whenever ddG is incorporated (shown in red), it causes termination of the growing chain. If dG (blue) is incorporated, the chain can continue growing. (B) When the sequencing reaction containing the ddG is run on a polyacrylamide gel, the fragments are separated by size. Each band directly represents a guanine in the original sequence.
B) Run on sequencing gel
Load sample — here
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