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Epigenetics clone


Visualization of histone acetylation

Histac for visualization of acetylation of the Histon

Catalog no. Name of clone Running title
RDB12840 Histac pcDNA3.1 For the detection of acetylation at the 5th and 8th lysine residues of histone H4 by the BRDT bromodomain.
RDB12841 Histac-K12 pcDNA3.1 For the detection of acetylation at the 12th lysine residues of histone H4 by the BRD2 bromodomain.
RDB14340 pcDNA3.1(+)-Histac-H3K9/K14 For the detection of acetylation at the 9th and 14th lysine residues of histone H3 by the BRD4 bromodomain.
The fluorescent probe which detects acetylation of the histones H3 and H4 – Histone modification is thought to be one of the mechanisms of epigenetic gene regulation. However, probably because of lack of measures to observe living specimen, spatiotemporal dynamics analysis is in little progress.

Here, we offer fluorescent probes “Histac” deposited by Dr. Kazuki Sasaki (RIKEN Brain Science Institute) to detect global hyperacetylation of histones in real-time living cells.

What is Histac?
Histac consists of an acetylation-binding domain, histones H3 and H4 as substrate, and the two fluorescent proteins.

Under the deacetylated conditions of the histon, CFP and Venus (YFP) in the Histac molecule are closed and yield a yellow fluorescence (emission, 535 nm) by fluorescence resonance energy transfer (FRET). On the other hand, under the acetylated conditions, separation of the fluorescence proteins occurs by the structural change caused by the binding of the bromodomain to histone in the Histac molecule, and yield a blue fluorescence (emission, 480 nm).

Observation of the acetylation of histone by the Histac requires no antibody reaction or fixation of cells. Histac allows successive observation of acetylation of the histone in the introduced living animal cells.

In the original papers (Ito, T., 2011 and Sasaki, K., 2009), the authors observed fluorescence intensity ratio between CFP versus Venus (480 nm / 535 nm) progressively from 0 to 180 minutes after treatment of histone deacetylase inhibitor, trichostatin A (TSA).
The authors proved that Histac as a powerful tool to show the dynamics of acetylation of histone in living cells.

Depositor: Dr. Kazuki Sasaki

References and Related Articles:

  • Nakaoka S, et al. A Genetically Encoded FRET Probe to Detect Intranucleosomal Histone H3K9 or H3K14 Acetylation Using BRD4, a BET Family Member. ACS Chem. Biol. 11 (3): 729-733, 2016.
  • Ito T, et al. Real-Time Imaging of Histone H4K12-Specific Acetylation Determines the Modes of Action of Histone Deacetylase and Bromodomain Inhibitors. Chemistry & Biology, 18(4):495-507, 2011.
  • Sasaki, K., et al. Real-time imaging of histone H4 hyperacetylation in living cells. PNAS, 106(38):16257-62, 2009.
  • RIKEN Research. Nov, 2009.
    URL : http://www.riken.jp/en/research/rikenresearch/highlights/6073

 

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Visualization of DNA methylation

pEGFP-MBD-nls for visualization of DNA mathylation

Catalog no. Name of clone Running title
RDB8344 pEGFP-MBD-nls Expression clone of methylated DNA binding domain (MBD) of human MBD1

To investigate relation between nuclear organization and epigenetic state of the genome, a strategy that allows simultaneous visualization of the epigenetic state such as methylation of DNA and nuclear structure has been required.

The expression clone pEGFP-MBD-nls allows you visualization of DNA methylation patterns in living cells.

The expression clone pEGFP-MBD-nls expresses an enhanced green fluorescent protein (EGFP) as a reporter. The reporter protein is fused with the methylated DNA binding domain (MBD) and the nuclear localization signal (nls) sequence of human methyl CpG-binding domain protein 1 (MBD1). The EGFP-MBD-nls protein recognizes the methylated DNA and you can follow status of the DNA methylation in situ under physiological conditions using the pEGFP-MBD-nls expression clone.

Depositor: Dr. Kuniya Abe

References and Related Articles:

 

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(GRP0030e 2012.11.07 T.M.)

2017.10.02