Yeast Histone Purification
Yeast recombinant histones are purified from a pET-based E. coli expression vector.
Contents
Section titled “Contents”1. Before You Begin
Section titled “1. Before You Begin”Urea and carbamylation of proteins: With heat and time, urea can break down into ammonium cyanate. Isocyanic acid will attack primary amines in proteins (Arg, Lys, N-terminus) to make a carbamylated protein. This interferes with histone structure and function. Many early or sloppy in vitro studies have suffered from this problem for decades. Source: http://www.ionsource.com/Card/carbam/carbam.htm
2. Purification
Section titled “2. Purification”- yeast H2A
- yeast H2B
- yeast H3
- yeast H4
3. Nucleosome Reconstitution
Section titled “3. Nucleosome Reconstitution”For now, read the methods paper (Methods Mol Biol vol. 778, p. 243).
4. Salt dialysis onto DNA
Section titled “4. Salt dialysis onto DNA”For now, read the methods paper (Methods Mol Biol vol. 778, p. 243).