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Sequencing Lec Notes
Cell Biology (BIO3153)
University of Ottawa
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Sequencing Lec Notes
DNA Sequencing When DNA polymerase catalyzes the formation of a phosphodiester link between a free 5-phosphate group on the sugar residue of a second nucleotide and a free 3 hydroxyl on the deoxyribose sugar moiety of one nucleotide, these are joined together.
Be that as it may, the artificial dideoxynucleotides ddATP, ddGTP, ddCTP, and ddTTP have no 3 - hydroxyl on their sugar buildup ,thus assuming they are integrated into a developing DNA chain, combination will stop. C 1′Each contains the four deoxynucleotides, DNA polymerase, the DNA template, and the primer.
Additionally, each of the mixtures has a low concentration of one of the four dideoxynucleotides—ddATP, ddGTP, ddCTP, or ddTTP—that sets it apart from the others. Let's examine what takes place in the tube depicted in Figure7 that contains ddTTP.
A respectable starting point that DNA polymerase experiences in the DNA format to be sequenced is an A. Since the cylinder contains considerably more dTTP than ddTTP, DNA polymerase will add a dTTP to most of the preliminary
particles. Notwithstanding, a little part of the groundworks will have a ddTTP added to them rather than dTTP since DNA polymerase can utilize either nucleotide as a substrate.
Since there is no OH group on the sugar's third carbon, DNA polymerase is unable to attach ddTTP to ddCTP. As a result, DNA synthesis is halted.
However, a dTTP was added to the majority of strands, allowing DNA polymerase to continue building the expanding strand. Similarly as in the past, these strands can develop no further. Each time an A appears on the template strand, this procedure will be repeated.
● At the point when the response is over, the cylinder will contain a combination of DNA pieces of various lengths, every one of which closes in addition to TTP. ● The portion of the tube containing ddTTP whose growth was halted after the first base, T, contains the smallest DNA molecules. ● The sequence of the template strand can be inferred from the fact that the new chain follows a sequence that is complementary to that of its template strand.
Sequencing Lec Notes
Course: Cell Biology (BIO3153)
University: University of Ottawa
