5 replies to this topic

[ramblingrat]

Hello everyone,

I am designing a project involving non-tumor DNA. We intend to do whole genome sequencing. So I was wondering  what would be the sample of choice? Buccal swab? Saliva? Chewing gum? Mouthwash?
Do I get enough material for next generation sequencing from these?
What are the considerations (smokers)?
Which are the pros and cons of there different methods (swab vs. saliva vs. other)?

Since I have no experience with NGS and isolation of DNA from such tissues any help/advise would be appreciated!! )

All the best,
the rat

[leelee]

Just a question- is the non tumour DNA that you are wanting to isolate and sequence to be used as a comparison for tumour DNA?

[ramblingrat]

hi leelee,

yes, it is and the tumor is leukemia, so it should not be anything using blood cells or being contaminated with it.

[leelee]

Why should it not be blood cells? I would have thought the best control for cancerous cells would be their non-cancerous counterpart, so you would actually want blood cells?

Am I missing something?

[ramblingrat]

Hey, thanks for your thoughts. Actually the microenvironment plays a major role in leukemias, therefore "non-tumor" blood cells are a bad choice because they are often mutated as well and consequently a relevant co-factor in tumor-progression.

[leelee]

Ah ok, that makes sense.

In that case, I think you could go with buccal cells. I know that when I teach undergrads and we isolate DNA in this way, we get plenty, so you should be able to get enough. We get them to scrape their inside cheek with a plastic spoon, then proceed to isolate the DNA from their. Its really gross, but effective