Hi everyone,
thank you in advance for your help.
Can isohelix genefix be used as a collection method to assess methylation levels in DNA from human saliva?
I.e. would it be ok to keep the saliva at room temperature in the hisohelix genefix buffer for several days? Will this affect the methylation status of the DNA or will DNA methylation be stable under these conditions?
Thank you immensely for your help,
Dr. Antonio Inserra
Depending on how its fixed in that buffer then yes, if the cells are kept in alive in suspension however there may be some change.
Your best bet though and to have an answer if asked when publishing it to run a control sample, same sample of cells from the same source (cultured or otherwise) and suspend them in the medium. Test one within a few hours and the other in several days or however long you think it will be for the real sample cohort. Then you can compare the results, run in triplicate or similar. You should have a good answer then
Dear Stephen, thank you very much for your answer.
What do you mean by "depending on how it is fixed in that buffer"? Samples have been collected, and stored in the buffer that comes with the kit. Unfortunately we are not able to run the control you suggest because samples have been collected in a remote location and will only be analysed later on in a different location.
Also, is DNA stored in this conditions viable for a genome-wide methylation study?
Thank you again.
With kind regards,
Antonio
DNA can be extracted from formalin fixed tissues and ethanol/methanol fixed tissues with no issue. This process essentially halts the replication machinery so there shouldnt be and more DNA alterations. I am not sure about the Isohelix buffer. Not sure what it is made of or for. Some buffers will preserve the cells by fixation essentially killing them in the process but others that would be more tailored for microbiology may preserve the cells in a live state. If the Isohelix preserves the cells live then there is a chance of alterations due to stress on the cell but if its fixed it should be fine.
Regarding the controls I would try get a Isohelix preservation buffer bottle and do it in the lab with a positive control if you have one or something similar. But yea I imagine it will be fine to run for whole genome analysis.
Best of luck with it
Stephen
Thanks immensely Steven. We are evaluating our possibilities
All the best
Antonio
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