Nucleosome, Recombinant Human, H3K14ac dNuc Description: Mononucleosomes assembled from recombinant human histones expressed in E. coli (two each of histones H2A, H2B, H3 and H4; accession numbers: H2A-P04908; H2B-O60814; H3.2-Q71DI3 *; H4-P62805) wrapped by 147 base pairs of 601 positioning sequence DNA. Histone H3 (created by a proprietary semi-synthetic method) contains acetyl-lysine at position 14. The nucleosome is the basic subunit of chromatin. The 601 sequence, identified by Lowary and Widom, is a 147-base pair sequence that has high affinity for histone octamers and is useful for nucleosome assembly and contains a 5' biotin-TEG group. * H3K14ac has a Cys to Ala substitution at position 110.
Nucleosome, Recombinant Human, H3K14ac dNuc Formulation: Nucleosome, Recombinant Human, H3K14ac (27.3 µg protein weight, 50 µg total weight) in 65 µl of 10 mM Tris HCl, pH 7.5, 25 mM NaCl, 1 mM EDTA, 2 mM DTT, 20% glycerol. Molarity = 3.84 μmolar. MW = 200,521 Da.
Nucleosome, Recombinant Human, H3K14ac dNuc Storage and Stability: Stable for six months at -80°C from date of receipt. For best results, aliquot and avoid multiple freeze/thaws.
Nucleosome, Recombinant Human, H3K14ac dNuc Application Notes: Nucleosome, Recombinant Human, H3K14ac dNucs are highly purified and are suitable for use as substrates in enzyme screening assays or for effector protein binding experiments. Nucleosome, Recombinant Human, H3K14ac from EpiCypher does not contain free DNA which could alter assayed activities.
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Western Blot Data: Western Analysis of Nucleosome, Recombinant Human, H3K14ac. Top Panel: Unmodified H3 (Lane 1) and H3K14ac containing nucleosomes (Lane 2) were probed with an anti-H3K14ac antibody and analyzed via ECL readout. Only the H3K14ac sample produced a detectable signal. Bottom Panel: Detail from Coomassie stained gel showing unmodified nucleosomes (Lane 1) and H3K14ac nucleosomes (Lane 2). (Click image to enlarge)
Mass Spec Data: Synthetic H3K14ac protein analyzed by high resolution mass spectrometry. Expected mass = 15266.8 Da. Determined mass = 15,265.5 Da. (Click image to enlarge)
Protein Gel Data: Coomassie stained PAGE gel of proteins in Nucleosome, Recombinant Human, H3K14ac (1 µg) to demonstrate the purity of the histones in the preparation. Sizes of molecular weight markers and positions of the core histones (H2A, H2B, H3K14ac and H4) are indicated. (Click image to enlarge)
DNA Gel Data: Nucleosome, Recombinant Human, H3K14ac run on a native PAGE gel and stained with ethidium bromide to visualize DNA. Lane 1: Free DNA extracted from nucleosomes (200 ng). Lane 2: Intact nucleosomes (400 ng). (Click image to enlarge)