Histone H3 and H4 were connected with 1 or 2 biotin through a short linker. And then they were stained to a dye through a streptavidin.
Just after the biotinylation, the standard SDS-PAGE gel cannot give desired bands at around 20 kDa.
- The band of biotinylized H3 is slightly more clear than the band of H4, but much lighter than the band of Histone without biotinylation.
- In the image of stained gel, stronger signal can be found in the top wells. So I assume it is aggregation of biotinylized hitone.
Any comment is welcomed!!!
if it is aggregation via hydrophobic interaction, try linking to a modified biotin with a polyethylene glycol chain instead, this will increase polarity and improve solubility
You mean you added Streptavidin and then analyzed the sample? If you did, take into account that if you don't boil the samples, the biotin-streptavidin interaction can survive through SDS-PAGE.
Hope it helps!
Dear Orenda Zhao,
can you provide details about the protocol used for biotinylation? Which strategy and reagent(s) have you used? Have you loaded the non-modified H3 and H4 for comparison?
Cheers Markus
- Badges
- Science method