Question

E.coli, to be replicated, is placed in a medium containing radioactive thymidine for five minutes. Then it is made to replicate in a normal medium. Which of the following observations will be correct?
(a) Both strands of DNA will be radioactive.
(b) One strand of DNA will be radioactive.
(c) Each strand of DNA will be half radioactive.
(d) None of the strands of the DNA will be radioactive.

Answer 1

Hint: The thymidine incorporation assay, the most common assay, uses a technique in which, during mitotic cell division, a radioactive nucleoside, $^3H$-thymidine, is inserted into new chromosomal DNA strands. The need to use radioisotopes has been replaced by more recently established assays.

Complete answer:
DNA replication is a semi-conservative process in which a strand of DNA serves as a blueprint for the synthesis of a new strand and generates two new molecules of DNA, each with a new strand and a parental strand. In lieu of a hydrogen atom in thymidine, coli with tritiated thymidine contains tritium. DNA isolated in both strands only after treatment and before cell division demonstrates radioactivity. This parental DNA replication of DNA with tritiated thymidine synthesizes the DNA strand of the daughter. The new DNA duplex is a combination of the parental strand with tritiated thymidine and a new DNA strand with normal thymidine since the daughter DNA strand carries normal thymidine. 50 percent normal DNA and 50 percent DNA intermediate DNA are generated by the second round of cell division. Replication of DNA does not split the strand of parental DNA.

Additional information: The benefit of these assays of incorporation is that they are direct proliferation measures. Indirect methods such as those that assess mitochondrial activity (e.g. MTT) need additional validation since, without affecting cell viability, test agents could influence the processing of MTT. The consumer should be conscious that it is not inherently proof of an anti-angiogenic effect to recognize test agents that inhibit proliferation. These agents can, in some cases, simply be toxic to the cell.
So, the correct answer is ‘(b) One strand of DNA will be radioactive’.

Note: To assess the degree of cell division that has occurred in response to a test agent, a scintillation beta-counter is used to measure the radioactivity in DNA retrieved from the cells. In the study of DNA synthesis in bacteria, radioactive labeling techniques are of significant importance. Radioactive thymine or thymidine is commonly used to mark DNA in Escherichia coli since these compounds are directly incorporated into DNA.