For human and amphibian cells I use Farnham Lysis Buffer with Protease inhibitors. After a 10 min incubation on ice I pipette up and down 20 times each sample with a 200 uL tip (note: cut 0.5 cm of the tip to make the hole a little bigger).

This is strange, your recipe is 100% correct

I think what Mr. gilberto suggested (Farnham buffer with PI) is only used when you want to extract protiens

However, try to prepare again whole recipe in smal amount and check again, otherwise order a commercial kit

Yes I tried changing all my solutions and I tested the pH but I think something is missing. You cannot dissociate tissues like you dissociate isolated cells.

Thanks anyway !

Yes I already tried increasing NP-40 concentration from 0,1 % to 1% and also adding Tween-20 and Digitonin but still nothing.

I agree with you that the protocol should be different and I saw the OMNI-seq alternative protocol. The only problem is that wing discs are really small tissues. I might use the option proposed for thin, frozen tissue sections but I still will have less material.

Thanks for your wishes !

What type of experiments did you perform than ? Because I want to avoid cell death not to induce a biais inside my results.

Actually I found a protocol that optimized tissues dissociation to minimize cell death (doi : 10.1080/19336934.2016.1173296 , A rapid, gentle and scalable method for dissociation and fluorescent sorting of imaginal disc cells for mRNA sequencing). I might try this one.

Fanny Chazal how did you solve your problem?

I'm trying to do Atac-seq too, and the buffer also didn't work for my cells (embryos). I'm considering the use of trypsin, but not sure if this would be the best approach.