Hello everyone,
I have done DNA sequencing on mouse samples after treatment with drug.. i wanted to see if there was any mutations caused by that drug.. I have found some mutations in P53 but unfortunately they do not have human homologues!
My question is, if they don't have human homologues does that mean they are not important!? is it possible that they only cause mutations in mouse yet not to humans? as the potential mutation was found in both treated and untreated tumors. the mouse did not bare that mutation prior to the experiment as genotyping was performed.
forgive me for my basic questions but this is my first time doing sequencing analysis.
I appreciate your answers
thank you
* how did you identify the mutations? This is important.
* what do you mean by "have human homologues"?
@shaojun Xie by sanger sequencing.. I have looked for the presence of two peaks for each nucleotide.
what i meant was that mutation is not found to be relevant to humans as seen from cosmic data base
Dear Deliah,
You may find this publication helpful.
p53 Mouse Models of Human Cancer By: Yvette Bren-Mattison, Ph.D. (MBC Pharma Inc.) © 2015 Nature Education 8(4):1
Clearly, mouse models were invaluable in helping researchers understand the complex functions and interactions of p53. Scientists have created similar models for many other cancer-related genes that have been very helpful in studies of human cancer biology.
However, most human disorders are polygenic, where a combination of alleles or genes are involved in disease. Mice that are genetically engineered usually target a single gene, thus being monogenic. There is a far greater genetic diversity in humans than in the inbred lines of mice used in the laboratory. The contribution of environmental factors is rarely considered when evaluating the phenotypes of genetically engineered mice. Some of these environmental factors include diet, exercise, exposure to pollutants, and stress — all of which are tightly controlled or absent in laboratory settings (Beckers et al. 2009).
Have you thought of inserting the human analogue into the mouse genome and observing any mutation in the presence of the drug?
Annwyne
@Annwyne Thank you so much .. could you se elaborate more about your last idea? did you mean inserting the human p53 gene into the mouse genome followed by the same treatment?
Many thanks
Although this in vivo procedure is not my area of expertise, I have engaged in in vitro investigations using lymphocytes, investigating genes and gene expression. As a medical educator, I have encountered and explained these methods in theory if not in practice. There are established methods for designing target vectors for inserting genes into embryonic stem cells. Cells containing the targeted vector into the targeted gene are selected and the target gene is knocked out in these cells. The gene is injected into the cells of the normal developing mouse embryo. The resulting homozygous chimeric mouse is bred and exposed to the drug of interest. A great deal of useful knowledge can be obtained from these procedures.
This method is described very well in the paper that I suggested in the previous message. (https://www.nature.com/scitable/nated/topicpage/p53-mouse-models-of-human-cancer-132732025)
https://www.nature.com/scitable/nated/topicpage/p53-mouse-models-of-human-cancer-132732025
Hi Delilah,
I would also suggest trying 3D modeling of the protein both with and without the mutation you are finding. A good (and free) place to start:
https://swissmodel.expasy.org/
In terms of whether your treatment is causing a mutation, if I understand you correctly it seems unlikely since you are finding the same mutation in both treated and untreated tumors. On the other hand, if the mutation is consistently found in tumor-cell DNA but not in the DNA of normal tissues, and it is in a critical gene like P53, it seems possible that it is an important mutation; but, this would require further validation of course.
Good luck!
Hi Deliah,
I agree with Matthew about proteomics as it is certainly a further step, after genomics and transcription studies but may prove more complicated. In the past these kind of studies depended on investigating mRNA expression. There are some very valuable findings from 3D proteomic analysis. Post-translational modifications to proteins can give a very much-improved picture of protein function. An exciting suggestion!
Best wishes Annwyne
- Badges
- Science topic
- Similar topics
- Chemistry
- Pharmacology
- Pharmaceuticals
- Drugs