Histones are the main constituents of the protein part of chromosomes of eukaryotic cells. They are rich in the amino acids arginine and lysine and have been greatly conserved during evolution. Histones pack the DNA into tight masses of chromatin. Two core histones of each class H2A, H2B, H3 and H4 assemble and are wrapped by 146 base pairs of DNA to form one octameric nucleosome. Histone tails undergo numerous post-translational modifications, which either directly or indirectly alter chromatin structure to facilitate transcriptional activation or repression or other nuclear processes. In addition to the genetic code, combinations of the different histone modifications reveal the so-called "histone code". Histone methylation and demethylation is dynamically regulated by respectively histone methyl transferases and histone demethylases. Acetylation of H4K20 is associated with gene activation.
H4K20ac Antibody - ChIP-seq Grade (sample size)
Monoclonal antibody raised in rabbit against histone H4 acetylated at Lys20 (H4K20ac), using a KLH-conjugated synthetic peptide.
| Lot | 001 |
|---|---|
| Concentration | 1 μg/μl |
| Species reactivity | Human. Other species not tested. |
| Type | Monoclonal, ChIP-grade, ChIP-seq grade, CUT&Tag-grade |
| Purity | Affinity purified monoclonal antibody |
| Host | Rabbit |
| Storage Conditions | Store at -20°C; for long storage, store at -80°C. Avoid multiple freeze-thaw cycles |
| Storage Buffer | PBS containing 50% glycerol, 1% BSA and 0.09% azide |
| Precautions | This product is for research use only. Not for use in diagnostic or therapeutic procedures. |
| Applications | Suggested dilution | References |
|---|---|---|
| ChIP/ChIP-seq * | 0.5 - 1 μg/ChIP | Fig 1, 2 |
| CUT&Tag | 1 μg | Fig 3 |
| Western Blotting | 1:1,000 | Fig 4 |
| Immunofluorescence | 1:500 | Fig 5 |
* Please note that the optimal antibody amount per IP should be determined by the end-user. We recommend testing 0.5-5 µg per IP.
- Validation data
Figure 1. ChIP results obtained with the monoclonal antibody directed against H4K20ac
ChIP assays were performed using human HeLa cells, the antibody against H4K20ac (cat. No. C15210008) and optimized PCR primer sets for qPCR. ChIP was performed with the iDeal ChIP-seq kit (cat. No. C01010055) on sheared chromatin from 1,000,000 cells. A titration of the antibody consisting of 0.5, 1, 2 and 5 µg per ChIP experiment was analysed. IgG (1 µg/IP) was used as negative IP control. QPCR was performed with primers for the active EIF4A2 and GAPDH genes, used as positive controls, and for the inactive MYOD1 gene and the Sat2 satellite repeat region used as negative controls. Figure 1 shows the recovery, expressed as a % of input (the relative amount of immunoprecipitated DNA compared to input DNA after qPCR analysis).
Figure 2. ChIP-seq results obtained with the monoclonal antibody directed against H4K20ac
ChIP was performed with 0.5 µg of the antibody against H4K20ac (cat. No. C15210008) on sheared chromatin from 1,000,000 HeLa cells using the iDeal ChIP-seq kit as described above. The IP’d DNA was subsequently analysed on an Illumina HiSeq 2000. Library preparation, cluster generation and sequencing were performed according to the manufacturer’s instructions. The 50 bp tags were aligned to the human genome using the BWA algorithm. Figure 2 shows the peak distribution along the complete sequence and a 600 kb region of the human X chromosome (figure 2A and B) and in two genomic regions surrounding the EIF4A2 and GAPDH positive control genes (figure 2C and D).
Figure 3. CUT&Tag results obtained with the antibody directed against H4K20ac
CUT&Tag was performed on 50,000 K562 cells using 1 µg of the antibody against H4K20ac (cat. No. C15210012) and the Universal CUT&Tag kit (cat. No. C01070024). The libraries were subsequently analysed on an Illumina NovaSeqX sequencer (2x150 bp paired-end reads) according to the manufacturer’s instructions. The tags were aligned to the human genome (hg38) using the BWA algorithm. Figure 3 shows the peak distribution along the complete sequence and a 1.5 Mb region of chromosome 1 (figure 3A and B) and in 2 genomic regions on chromosome 12 and 7 (figure 3C and D, respectively).
Figure 4. CUT&Tag results obtained with the antibody directed against H4K20ac
CUT&Tag was performed on 50,000 K562 cells using 1 µg of the antibody against H4K20ac (cat. No. C15210012) and the Universal CUT&Tag kit (cat. No. C01070024). The libraries were subsequently analysed on an Illumina NovaSeqX sequencer (2x150 bp paired-end reads) according to the manufacturer’s instructions. The tags were aligned to the human genome (hg38) using the BWA algorithm. Figure 3 shows the peak distribution along the complete sequence and a 1.5 Mb region of chromosome 1 (figure 3A and B) and in 2 genomic regions on chromosome 12 and 7 (figure 3C and D, respectively).
Figure 5. Immunofluorescence using the monoclonal antibody directed against H4K20ac
HeLa cells treated with sodium butyrate were stained with the antibody against H4K20ac (Cat. No. C15210008, red) diluted 1:500. Actin filaments were stained with fluorescein phalladoin (green). - 出版物
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