Agrisera/PsbA | D1 protein of PSII, C-terminal (rabbit)/AS05 084/-蚂蚁淘生物


product information
Background		The psbA gene has been cloned from many species of plants, green algae, and cyanobacteria. The psbA gene is located in the chloroplast genome and encodes for the D1 protein, a core component of Photosystem II. PsbA/D1 is rapidly cycled under illumination in all oxygenic photobionts. Tracking PsbA pools using the Global PsbA antibody can show the functional content of Photosystem II in a wide range of samples. Alternative names: 32 kDa thylakoid membrane protein, photosystem II protein D1
Immunogen		KLH-conjugated synthetic peptide derived from available plant, algal and cyanobacterial PsbA sequences, including Arabidopsis thaliana UniProt: A4QJR4, TAIR: AtCg00020 , Oryza sativa P0C434, Populus alba Q14FH6, Physcomitrella patens Q6YXN7, Chlamydomonas reinhardtii P07753, Synechocystis sp. P14660 and many others
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		Western blot (WB)
Related products		AS01 016S \| PsbA protein standard for a quantitative western blotAS05 084PRE \| PsbA \| D1 protein of PSII, C-terminal , pre-immune serumAS11 1786 \| anti-PsbA N-terminal, rabbit antibodyAS10 704 \| PsbA \| D1 protein of PSII, DE-loop, rabbit antibodyAS13 2669 \| PsbA \| D1 protein of PSII, phosphorylated, rabbit antibody recommended secondary antibody Plant and algal protein extraction buffer Secondary antibodies
Additional information		Due to biology of PsbA (D1) protein a number of degradation products can apprear in a sample and may be observed when using anti-PsbA antibodies, including products having apparent molecular weights of 24kDa and 16kDa. D1 degradation is a complex set of events and the products observed can be influenced by both the extraction procedure and the physiology of the cells prior to harvest. Third, cross-linking may occur between D1 and cytochrome b559, shifting the protein higher in the gel. In cyanobacteria (PCC7942), three different bands were competed out by preincubating the antibody with the PsbA free peptide, indicating that all bands are indeed PsbA and its precursors or breakdown products. Competition assays were also performed with spinach and Chlamydomonas, confirming the identity of PsbA bands. Anti-PsbA antibodies will not detect D2 protein, as the peptide used to generate PsbA antibodies has no homology to the D2 sequence.

application information
Recommended dilution		1: 10 000 (WB)
Expected \| apparent MW		38 \| 28-30 kDa
Confirmed reactivity		Anabaena 7120, Arabidopsis thaliana, Artemisia annua, Arundo sp., Chlamydomonas reinhardtii, Chlorella ohadii, Chromera velia, Colobanthus quitensis Kunt Bartl, Coscinodiscus wailesii, Craterostigma sp., Cyanidioschyzon merolae, Cytisus cantabricus (Wilk.) Rchb. F, Ditylum brightwellii, Eucalyptus globulus, Glycine max, Hieracium pilosella L., Hordeum vulgare, Lasallia hispanica, Lindernia sp., Marchantia polymorpha (liverwort), Miscanthus x giganteus, Microcystis aeruginosa, Nicotiana benthamiana, Panicum miliaceum, Panax ginseng, Panicum maximum, Paulinella chromatophora (amoeba), Pheodactylum tricornutum CCAP 1055/1, Physcomitrella patens, Pinus strobus, Prochlorococcus sp. (surface and deep water ecotype), Spartina alterniflora, Spirodela polyrhiza, Symbiodinium sp, Synechococcus sp. PCC 7942, Syntrichia muralis, Triticum aestivum, Zea mays
Predicted reactivity		Lycopersicum esculentum, Medicago sativa, Pisum sativum, Sesamum indicum, and other di and monocots,conifers, brown and red algae, cyanobacteria; cellular [compartment marker] of thylakoid membrane
Not reactive in		no confirmed exceptions from predicted reactivity known in the moment
Additional information		The antibody is appropriate for detecting both, 24 kDa or the 10 kDa C-terminal fragments, whichever is generated under given treatment conditions. In our analysis we have seen both, ca. 24 kDa and ca. 10 kDa fragments from different samples, depending on treatments and isolation procedures. Rabbit anti-PsbA antibody can detect more than one band of PsbA protein, e.g. precursor and mature protein as compare to the hen anti-PsbA antibodies AS01 016. This antibody will detect the phosphorylated form of D1 as an alternate band to the main band on a high resolution gel.The antibody will bind to cross-linked proteins: D1/D2, D1/cyt b559, D1/CP43.
Selected references		Ananyev et al. (2017). Photosystem II-Cyclic Electron Flow Powers Exceptional Photoprotection and Record Growth in the Microalga Chlorella ohadii.Biochim Biophys Acta. 2017 Jul 19. pii: S0005-2728(17)30105-6. doi: 10.1016/j.bbabio.2017.07.001. Sharwood et al. (2017). Linking photosynthesis and leaf N allocation under future elevated CO2 and climate warming in Eucalyptus globulus. J Exp Bot. 2017 Feb 1;68(5):1157-1167. doi: 10.1093/jxb/erw484. Míguez et al. (2017). Diversity of winter photoinhibitory responses: A case study in co-occurring lichens, mosses, herbs and woody plants from subalpine environments. Physiol Plant. 2017 Feb 14. doi: 10.1111/ppl.12551. Romanowska et al. (2017). Differences in photosynthetic responses of NADP-ME type C4 species to high light. Planta. 2017 Mar;245(3):641-657. doi: 10.1007/s00425-016-2632-1. Epub 2016 Dec 18. Yang-Er Chen et al. (2017). Responses of photosystem II and antioxidative systems to high light and high temperature co-stress in wheat. J. of Exp. Botany, Volume 135, March 2017, Pages 45–55. Jallet et al. (2016). Photosynthetic physiology and biomass partitioning in the model diatom Phaeodactylum tricornutum grown in a sinusoidal light regime. Algal research, doi:10.1016/j.algal.2016.05.014. Heinnickel et al. (2016). Tetratricopeptide repeat protein protects photosystem I from oxidative disruption during assembly. Proc Natl Acad Sci U S A. 2016 Mar 8;113(10):2774-9. doi: 10.1073/pnas.1524040113. Treves et al. (2016). The mechanisms whereby the green alga Chlorella ohadii, isolated from desert soil crust, exhibits unparalleled photodamage resistance. New Phytol. 2016 Feb 8. doi: 10.1111/nph.13870 Wang et al. (2015). The combined effects of UV-C radiation and H2O2 on Microcystis aeruginosa, a bloom-forming cyanobacterium. Chemosphere. 2015 Jun 16;141:34-43. doi: 10.1016/j.chemosphere.2015.06.020. Bancel et al. (2015). Proteomic Approach to Identify Nuclear Proteins in Wheat Grain. J Proteome Res. 2015 Sep 8. Vandenhecke et al. (2015). Changes in the Rubisco to photosystem ratio dominates photoacclimation across phytoplankton taxa. Photosynth Res. 2015 Jun;124(3):275-91. doi: 10.1007/s11120-015-0137-6. Epub 2015 Apr 11. Charuvi et al. (2015). Photoprotection Conferred by Changes in Photosynthetic Protein Levels and Organization during Dehydration of a Homoiochlorophyllous Resurrection Plant. Plant Physiol. 2015 Apr;167(4):1554-65. doi: 10.1104/pp.114.255794. Spence et al. (2014). Transcriptional responses indicate maintenance of photosynthetic proteins as key to the exceptional chilling tolerance of C4 photosynthesis in Miscanthus × giganteus. J Exp Bot. 2014 Jul;65(13):3737-47. doi: 10.1093/jxb/eru209. Epub 2014 Jun 22. Pandey and Pandey-Rai (2014). Modulations of physiological responses and possible involvement of defense-related secondary metabolites in acclimation of Artemisia annua L. against short-term UV-B radiation. Planta. 2014 Jul 15. Vinyard et al. (2014). Engineered Photosystem II reaction centers optimize photochemistry vs. photoprotection at different solar intensities. J Am Chem Soc. 2014 Mar 3. Malnoë et al. (2014). Thylakoid FtsH Protease Contributes to Photosystem II and Cytochrome b6f Remodeling in Chlamydomonas reinhardtii under Stress Conditions. Plant Cell, Jan 21.

application example 1

2 µg of total protein from (1) Arabidopsis thaliana leaf extracted with Protein Extration Buffer, PEB (AS08 300), (2) Hordeum vulgare leaf extracted with PEB, (3) Chlamydomonas reinhardtii total cell extracted with PEB, (4) Synechococcus sp. 7942 total cell extracted with PEB, (5) Anabaena sp. total cell extracted with PEB were separated on 4-12% NuPage (Invitrogen) LDS-PAGE and blotted 1h to PVDF. Blots were blocked immediately following transfer in 2% ECL Advance blocking reagent (GE Healthcare) in 20 mM Tris, 137 mM sodium chloride pH 7.6 with 0.1% (v/v) Tween-20 (TBS-T) for 1h at room temperature with agitation. Blots were incubated in the primary antibody at a dilution of 1: 50 000 for 1h at room temperature with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, then washed once for 15 min and 3 times for 5 min in TBS-T at room temperature with agitation. Blots were incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, recommended secondary antibody AS09 602) diluted to 1:50 000 in 2% ECL Advance blocking solution for 1h at room temperature with agitation. The blots were washed as above and developed for 5 min with ECL Advance detection reagent according the manufacturers instructions. Images of the blots were obtained using a CCD imager (FluorSMax, Bio-Rad) and Quantity One software (Bio-Rad).

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application example 2

Varying amounts of PsbA protein standard (AS01 016S) 250 fmol (1), 125 fmol (2), 62.5 fmol (3), 31.25 fmol (4), 15.625 fmol (5) and 2 µg of total protein from Med4 (6,7) extracted with Protein Extration Buffer, PEB (AS08 300). Samples were diluted with 1X sample buffer (NuPAGE LDS sample buffer (Invitrogen) supplemented with 50 mM DTT and heat at 70°C for 5 min and keept on ice before loading. Protein samples were separated on 4-12% Bolt Plus gels, LDS-PAGE and blotted for 70 minutes to PVDF using tank transfer. Blots were blocked immediately following transfer in 2% blocking reagent (GE RPN 2125; Healthcare) or 5% non-fat milk dissolved in 20 mM Tris, 137 mM sodium chloride pH 7.6 with 0.1% (v/v) Tween-20 (TBS-T) for 1h at room temperature with agitation. Blots were incubated in the primary antibody at a dilution of 1: 10 000 (in blocking reagent) for 1h at room temperature with agitation. The antibody solution was decanted and the blot was rinsed briefly twice, and then washed 1x15 min and 3x5 min with TBS-T at room temperature with agitation. Blots were incubated in secondary antibody (goat anti-rabbit IgG horse radish peroxidase conjugated, recommended secondary antibody AS09 602, Agrisera) diluted to 1:25 000 in blocking reagent for 1h at room temperature with agitation. The blots were washed as above. The blot was developed for 5 min with TMA-6 (Lumigen) detection reagent according the manufacturers instructions. Images of the blots were obtained using a CCD imager (VersaDoc MP 4000) and Quantity One software (Bio-Rad). Exposure time was 30 seconds.

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