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H3 | Histone H3 (polyclonal, serum)

265 €
Buy 2 items of this product for 198 €/each
Buy 3 items of this product for 180 €/each

AS10 710 | clonality: polyclonal | host: rabbit | reactivity: A. thaliana, S. lycopersicum, V. faba, P. patens, S. europaea, Z. mays, Ch. reinhardtii | cellular [compartment marker] of nucleoplasm

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AS10 710

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product information
Background

Histone 3 (H3) located in nuclei, incorporated into chromatin. Present in nucleosome together with H2A, H2B and H4.

Immunogen

KLH-conjugated synthetic peptide derived from known H3 sequences, inluding Arabidopsis thaliana H3.3 P59169 (At4g40030, At4g40040, At5g10980), H3.2 P59226 (At1g09200, At3g27360, At5g10390, At5g10400, At5g65360), H3-like 2 Q9FXI7 (At1g19890)

Host Rabbit
Clonality Polyclonal
Clone
Purity Serum
Format Lyophilized
Quantity 50 µl
Reconstitution For reconstitution add 50 µl of sterile water.
Storage

Store lyophilized/reconstituted at -20°C; once reconstituted make aliquots to avoid repeated freeze-thaw cycles. Please, remember to spin tubes briefly prior to opening them to avoid any losses that might occur from lyophilized material adhering to the cap or sides of the tubes.

Tested applications
ChIp-qPCR (ChIp-qPCR), Immunocytochemistry (ICC), Western Blot (WB)
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AS10 710PRE | H3 | histone H3, pre-immune serum

collection of antibodies to DNA/RNA/cell cycle


Plant and algal protein extraction buffer

Secondary antibodies

Additional information

Cellular [compartment marker] of nucleoplasm, loading control antibody for Chlamydomonas reinhardtii

application information
Recommended dilution

1 : 5000 with standard ECL (WB), 1: 100 - 1: 500 (ICC), 2 µl of antibody/500 µl solution (ChIp-qPCR)

Expected | apparent MW

15 | 17 kDa

Confirmed reactivity

Arabidopsis thaliana, Brassica oleracea, Capsicum annuum, Chlamydomonas acidophila, Chlamydomonas reinhardtii, Physcomitrella patens, Salicornia europaea, Solanum lycopersicum, Solanum sogarandinum, Solanum tuberosum, Vicia faba, Zea mays

Predicted reactivity

Brachypodium distachyon, Brassica napus, Hordeum vulgare, Nicotiana tabacum, Malus domestica, Medicago sativa, Triticum aestivum, Pinus pinaster, Pisum sativum, Oryza sativa, Zea mays, Vitis vinifera, Volvox sp.

Not reactive in

No confirmed exceptions from predicted reactivity known in the moment

Additional information
This product can be sold containing ProClin if requested

Protocol for isolation of cytosolic and nuclear fractions can be found here.
Specific fluorescence in ICC has been observed for interphase nuclei as well as around centromer region (where Ser10 of histone H3 is phosphorylated) in mitotic chromosomes.
Selected references Correa-Galvis et al. (2016). Photosystem II Subunit PsbS Is Involved in the Induction of LHCSR Protein-dependent Energy Dissipation in Chlamydomonas reinhardtii. J Biol Chem. 2016 Aug 12;291(33):17478-87. doi: 10.1074/jbc.M116.737312.
Castellano et al. (2016). A pathogenic long noncoding RNA redesigns the epigenetic landscape of the infected cells by subverting host Histone Deacetylase 6 activity. New Phytol. 2016 Sep;211(4):1311-22. doi: 10.1111/nph.14001. Epub 2016 May 12.
Ghandi et al. (2016). Tomato yellow leaf curl virus infection mitigates the heat stress response of plants grown at high temperature. Sci Rep. 2016 Jan 21;6:19715. doi: 10.1038/srep19715.
Gorovits et al. (2016). Tomato yellow leaf curl virus confronts host degradation by sheltering in small/midsized protein aggregates. Virus Res. 2016 Feb 2;213:304-13. doi: 10.1016/j.virusres.2015.11.020. Epub 2015 Dec 1
Li et al. (2015). Three SAUR proteins SAUR76, SAUR77 and SAUR78 promote plant growth in Arabidopsis. Sci Rep. 2015 Jul 24;5:12477. doi: 10.1038/srep12477.
Spijkerman et al. (2014). CO2 acquisition in Chlamydomonas acidophila is influenced mainly by CO2, not phosphorus, availability. Photosynth Res. 2014 Sep;121(2-3):213-21. doi: 10.1007/s11120-014-0016-6. Epub 2014 Jun 7.
Rihan et al. (2014). The effect of molybdenum on the molecular control of cold tolerance in cauliflower (Brassica oleracea var. botrytis) artificial seeds. Plant Cell, Tissue and Organ Culture (PCTOC) April 2014
Szabala
et al. (2014). Accumulation of acidic SK3 dehydrins in phloem cells of cold- and drought-stressed plants of the Solanaceae. Planta, Jan 7.
Wobbe and Nixon
(2013). The mTERF protein MOC1 terminates mitochondrial DNA transcription in the unicellular green alga Chlamydomonas reinhardtii. Nucleic Acids Res. May 6.

application example

 Histone H3 plant antibody western blot example

1.2 μg of Arabidopsis thaliana chromatin-enriched fraction (1) and 3.75 µg of total protein from 4-weeks-old Arabidopsis thaliana leaves (2), and were separated on 12% SDS-PAGE and blotted 50 mins to Immobilon-P (Millipore, semi-dry) PVDF membrane. Blots were blocked immediately following transfer in MTBS-T (5% milk) for 30 mins at room temperature with agitation. Blots were incubated in the primary antibody at a dilution of 1:5000 for 1h at room temperature with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed 3 times for 3 min in TBS-T at room temperature with agitation. Blots were incubated in secondary antibody (anti- IgG horse radish peroxidase conjugated, from Agrisera, AS09 602) diluted to 1:20 000 for 30 mins at room temperature with agitation. The blots were washed as above and developed for 5 min with ECL detection reagent (Roche) according to the manufacturers instructions. Exposure time was 30 seconds. Double band in chromatine-enriched fraction (1) has been outcompeted in peptide neutralization assay by peptide used to elicit H3 antibodies. Chromatin izolation was carried out as described (Zilberman et al. 2008) with minor modifications.

Courtesy of Weronika Sura and Dr. Piotr A. Ziolkowski (Department of Biotechnology, Adam Mickiewicz University, Poznan, Poland)

 

 

western

30 μg of 5 µl of Chlamydomonas reinhardtii protein saturated in 8M urea were separated on 15% SDS-PAGE and blotted for 1hour to 0.2 µm nitrocellulose at 100V using wet transfer system. Blots were blocked with 0.5% cold fish gelatin for 1hr at room temp with agitation. Blot was incubated in the primary antibody (anti-H3) at a dilution of 1:2500 for an hour at RT with agitation. The blots were washed with 3X 15min TBS-TT at RT with agitation. Blots as incubated in the secondary antibody (DayLight 800) 1:5000 dilution for 30min at RT with agitation and washed 1X with TBSTT for 15min, 1X with TBST for 15min before scanning with the ODyssey IRD scanner.

Courtesy of Dr. Betty Chung, University of Cambridge, United Kingdom
western blot using anti-H3 antibodies on tomato samples
5 µl of 15μg/µl Solanum lycopersicum protein saturated in 8M urea were separated on 15% SDS-PAGE and blotted for 1hour to 0.2 µm nitrocellulose at 100V using wet transfer system. Blots were blocked with 0.5% cold fish gelatin for 1hr at room temp with agitation. Blot was incubated in the primary antibody (anti-H3) at a dilution of 1:2500 for an hour at RT with agitation. The blots were washed with 3X 15min TBS-TT at RT with agitation. Blots as incubated in the secondary antibody (DayLight 800) 1:5000 dilution for 30min at RT with agitation and washed 1X with TBSTT for 15min, 1X with TBST for 15min before scanning with the ODyssey IRD scanner.

Courtesy of Dr. Betty Chung and Dr Zhengming Wang, University of Cambridge, United Kingdom





ICC

Immunocytochemical assays were performed according to the method described earlier (Rybaczek and Maszewski 2006). Excised apical parts of Vicia faba roots (1.5 mm long) were fixed for 45 min (18°C) in PBS-buffered 3.7% paraformaldehyde, washed several times with PBS and placed in a citric acid-buffered digestion solution (pH 5.0; 37°C for 45 min) containing 2.5% pectinase (Fluka), 2.5% cellulase (Onozuka R-10; Serva) and 2.5% pectoliase (ICN). After removing the digestion solution, root tips were washed 3 times in PBS, rinsed with distilled water and squashed onto Super Frost Plus glass slides (Menzel-Gläser). Air-dried slides were pretreated with PBS-buffered 5% BSA at 20°C for 50 min and incubated overnight in a humidified atmosphere (4°C) with rabbit antibody raised against H3 histone (Agrisera), dissolved in PBS containing 1% BSA (at a dilution of 1:50). Following incubation, slides were washed 3 times with PBS and incubated for 1 h (18°C) with Agrisera secondary goat anti-rabbit IgG DyLight®488 antibody (AS09 633, 1:1000). Nuclear DNA was stained with 4’,6-diamidino-2-phenyl-indole (DAPI, 0.4 μg/ml; Sigma-Aldrich). Following washing with PBS, slides were air dried and embedded in Vectashield Mounting Media for Fluorescence (Vector Laboratories). Observations were made using Optiphot-2 fluorescence microscope (Nikon) equipped with B-2A filter (blue light; λ ≈ 495 nm) for DyLight-conjugated antibodies and UV-2A filter (UV light; λ ≈ 365 nm) for DAPI. All images were recorded at exactly the same time of integration using DXM 1200 CCD camera.

Courtesy Dr. Dorota Rybaczek, Lodz University, Poland


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