i fractionated cell culture via a kit
but i do not want to use the kit further after cytoplasm removal because i am afraid the detergents for nuclear extraction might ruin protein protein interaction.
so i used buffer h and sonication, but the normal level of sonication for whole cells (level = 7) did not effect my sample and i still see the lump of DNA stuck together.
which level of sonication should i use?
I don't think sonication alone will separate the nucleic material. If you do get the power high enough to break up the lumps, it will almost certainly break the chains. Sorry, I realise that is not very helpful.
I will go with Andrew Kirkham, ultrasonication alone cannot able to break the nuclei membrane. when you are going for more power or Hz it will produce heat which may damage your nuclei material. My suggestion is to go with best kit method which doesnt ruin protein protein interaction. All the best
Thank you both for the helpful answers!
I wish to avoid the kit method is the detergents... do you have any experience with that?
If you use a DNase, that will help break up the nuclear material. If you're doing proteomics, benzinase (which digests DNA and RNA) can clean up your preps.
- Badges
- Science method