Hello,
To isolate nuclear proteins from cytoplasmic and mitochondrial proteins, we aim to use the Nuclear Exraction Kit from ActiveMotif (https://www.activemotif.com/catalog/100/nuclear-extract-kit). They showed with WB that they could isolate from HeLa cells the cytoplasmic extract, the nuclear extract and the nuclear pellet from the nuclear extract.
They used RNA-pol II as a nuclear marker : found in the nuclear pellet but not much in the nuclear extract (as showed below)
I have difficulties to understand what is the nuclear pellet ? Why RNA-pol II is found in the nuclear pellet but not in the nuclear extract ? What is the nuclear extract in that case ?
I thank you in advance for your help
RNA Pol II is DNA bound and the chromatin is going to spin out into the pellet from the nuclear fraction. DNA bound proteins are difficult to solubilize in most standard lysis buffers and require detergent or a denaturing agent to help get these proteins into the soluble fraction.
- Badges
- Science method