Question 1 Nucleic acid chemistry
Use the structures below to answer question 1 i – iii
i) Which structure (A – E) contains only a purine base?
ii) Which structure (A – E) contains only a nucleoside?
iii) Which structure (A – E) is the substrate for Reverse transcriptase? Question 1 continued
Use the structure below to answer questions 1 iv to ix Question 1 continued
Identify the following in the pentanucleotide strand on page 3:
iv) label the 3’ OH
v) circle a phosphodiester bond vi) identify a nitrogen atom in one of the
heterocyclic base rings with pKa ~4 vii) If this strand were copied by
Primase what would be the sequence of the product? Sequences are quoted
5’ to 3’
viii) How many hydrogen bonds would the double stranded oligonucleotide
produced in (vii) contain?
ix) If this was a part of a growing nucleotide chain, show where the next nucleotide
would add. What other product(s) would result?
Nucleic acid properties exploited in Molecular Biology
You have a dye, Sybr GreenI, which binds only to double stranded DNA (not single
stranded). Once bound, it fluoresces strongly and can be used to monitor DNA melting
(transition from double stranded to single stranded).
(i) On the axes provided draw the DNA melt curve for a sample of E. coli DNA which
has been mixed with Sybr GreenI.
On the same graph show the melt curve for a sample of DNA:
(ii) with a higher %(G+C) content,
(iii) from the same source (E coli) dissolved in a buffer with lower [Na+
(iv) from the same source (E coli) dissolved in a buffer at pH>9 for 30 min prior to
melting Make sure you label each curve.
A double stranded DNA solution with an absorbance of 1.0 at 260 nm has a concentration of
50 µg/ml while a single stranded DNA solution with the same absorbance would only have a
concentration of 35 µg/mL. Account for the difference in the absorption properties.
Explain how the twist in DNA arises.
Below is the structure of an antiretroviral drug, AZT, which is a modified nucleotide.
(i) What nucleoside/nucleotide does this drug “mimic”?
(ii) What structural feature of the nucleotide/nucleoside has been modified?
(iii) This drug will be taken up by the cell and rapidly phosphorylated to the nucleotide
triphosphate. As a nucleotide triphosphate what cellular process might this
modification interfere with and why?
(iv) What retroviral enzyme is it targeting and why does it have less effect on host cell
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A double stranded DNA solution with an absorbance of 1.0 at 260 nm has a concentration of 50 μg/ml while a single stranded DNA solution with the same absorbance would only have a concentration of 35 μg/mL. Account for the difference in the absorption properties.
The purine and pyrimidine bases in DNA show absorbance at 260 nm. In double stranded DNA, purine and pyrimidine bases are well stacked and they absrobs less UV due to better stacking interaction. But, this base stacking interaction is absent in case of single stranded DNA and they absobs more strongly than double stranded DNA.
Thus, an absorbance of 1.0 at 260 nm can be obtained by a concentration of 50 μg/ml of double stranded DNA, whereas a less amount of 35 μg/mL single stranded DNA can absorb the same amount of UV.
Explain how the twist in DNA arises.
Sugar and phosphate of the DNA are hydrophilic whereas, nitrogen bases are hydrophobic. Thus N- bases tends to remain in the core region of the double stranded DNA axis. Moreover, N-bases of two strands attracts each other while another force, the base stacking force attracts the bases of the same DNA strand above or below of that particular base. This happens in case of each and every base of the double stranded DNA molecule and to make balance between these two attractive forces along with hydrophobic interaction, double stranded DNA get twisted....