Question 2 It is known that some tumors are associated with tumor viruses. It re

Question

Question 2 It is known that some tumors are associated with tumor viruses. It remains unknown, however, whether or not other tumors such as those induced by chemical carcinogens are also associated with tumor viruses. Design a series of experiments to test the hypothesis that non-viral carcinogen-induced tumor cells contain a gene(s) that is altered (as compared to its unaltered state in the normal cell counterparts) and is responsible for the transformed phenotypes of the tumor cells. Include in your experimental design the following: 1) the overall strategy, 2) the choices of reagents including controls and the rationales behind each of the choices, 3) at least four types of transformation assays, including both in vitro and in vivo assays, to be used to measure the properties of transformed cells, and 4) the strategy to interpret the results and to draw a conclusion. Question 2 It is known that some tumors are associated with tumor viruses. It remains unknown, however, whether or not other tumors such as those induced by chemical carcinogens are also associated with tumor viruses. Design a series of experiments to test the hypothesis that non-viral carcinogen-induced tumor cells contain a gene(s) that is altered (as compared to its unaltered state in the normal cell counterparts) and is responsible for the transformed phenotypes of the tumor cells. Include in your experimental design the following: 1) the overall strategy, 2) the choices of reagents including controls and the rationales behind each of the choices, 3) at least four types of transformation assays, including both in vitro and in vivo assays, to be used to measure the properties of transformed cells, and 4) the strategy to interpret the results and to draw a conclusion. Question 2 It is known that some tumors are associated with tumor viruses. It remains unknown, however, whether or not other tumors such as those induced by chemical carcinogens are also associated with tumor viruses. Design a series of experiments to test the hypothesis that non-viral carcinogen-induced tumor cells contain a gene(s) that is altered (as compared to its unaltered state in the normal cell counterparts) and is responsible for the transformed phenotypes of the tumor cells. Include in your experimental design the following: 1) the overall strategy, 2) the choices of reagents including controls and the rationales behind each of the choices, 3) at least four types of transformation assays, including both in vitro and in vivo assays, to be used to measure the properties of transformed cells, and 4) the strategy to interpret the results and to draw a conclusion.

Explanation / Answer

Let us take the effect of chronic exposure of aflatoxin B1 (from Aspergillus spore) on liver cancer. It is known that aflatoxin induces p53 (a tumour suppressor) mutations leading to abrogation of its activit and initiation followed by progression of the tumour.

To detect whether the carcinogen has caused mutation in say p53, one can obtain p53 from normal liver cell line (AML-12) and obtain the p53 gene after cDNA synthesis and send it for sequencing to confirm the same. One should confirm the same in some other cell lines as well.

Once it is confirmed that aflatoxin has induced gene mutation, one can go for various functional studies and compare the effect of mutant and wild type p53. The assays are as follows:

1. Aflatoxin treated and untreated on liver cell line/liver cancer cell line, look for p53 increased or reduced downstream signalling events (using western blot, fluorescence microscopy).

2. One can look at the promotor activity difference between wild type and mutant.

3. One can also perform microsphere assay, migration assay to compare the effect with and without aflatoxin.

4. One can inject aflatoxin on mice and look at liver biopsy for various markers (including p53), perform immunohistochemistry of liver to compare the difference between untreated and treated mice. Also one can isolate liver cells from these treated mice and utilise flow cytometry to find out the difference between some of the well known receptors that get affected when treated with aflatoxin. One can also look at the differences in cell division compared to control using various cell division markers (flow cytometry or western blot) including the differences in G1/S/G2/M phases. The same can be done over a cell line as well. One should always confirm the mutation upon treatment before proceeding for each experiments.

These assays will definitely help any cancer biologists to look at the effect that the molecule causes on cancer by altering a particular gene.

One can also study structural aspects in terms of differences in wild type and mutant gene stabilities, in vitro activity etc.

Related Questions

Navigate

• Browse (All)
• Browse Q
• Subjects

---

---

Integrity-first tutoring: explanations and feedback only — we do not complete graded work. Learn more.