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UBQ11 | Ubiquitin (affinity purified)

AS08 307A  |  Clonality: Polyclonal  |  Host: Rabbit  |  Reactivity: A.thaliana, H.vulgare, L. angustifolius, M.domestica, N. benthamina, O. sativa, P. sativum, S.tuberosum, Z.mays

UBQ11 | Ubiquitin (affinity purified) in the group Antibodies for Plant/Algal / Developmental Biology / Signal transduction at Agrisera AB (Antibodies for research) (AS08 307A)

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product information
Background

Ubiquitin is a highly conserved regulatory protein expressed in all eukaryotic tissues. Originally this protein was called: Ubiquitous Immunopoietic Polypeptide. Its function is labeling of proteins for degradation through ubiquitin proteasome system (UPS).

Immunogen

Recombinant Arabidopsis thaliana ubiquitin, ubq11, with N-terminal His-tag; TAIR:  At4g05050

Host Rabbit
Clonality Polyclonal
Clone
Purity Affinity purified serum in PBS pH 7.4.
Format Lyophilized
Quantity 50 µg
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
AS08 307 | UBQ11 | Ubiquitin, rabbit antibody, provided in a serum format

AS10 307S | UBQ11 | Ubiquitin protein | positive control
Additional information
application information
Recommended dilution 1 : 10 000 (WB)
Expected | apparent MW

8.5 kDa

Confirmed reactivity Arabidopsis thaliana, Hordeum vulgare, Lupinus angustifolius, Malus domestica, Nicotiana benthamina, Oryza sativa, Pisum sativum, Solanum tuberosum, Zea mays
Predicted reactivity Brachypodium distachyon, Brassica napus, Capsella rubella, Cochliobolus heterostrophus, Citrus clementina, Citrus sinensis, Coffea canephora, Cymbidium faberi, Eucalyptus grandis, Erythrane guttata, Glycine max, Glycine soja, Ipomoea batatas, Medicago truncatula, Magnaporthe oryzae, Nicotiana tabacum, Phaseolus vulgaris, Populus trichocarpa, Prunus persica, Pyrus communis, Ricinus communis, Solanum nigrum,  Sorghum bicolor, Triticum aestivum, Theobroma cacao, Vitis vinifera
Not reactive in Algae
Additional information

Antibody can be used to check qubiquitination status in the whole plant extracts.

Technical note: It is very difficult to detect ubiquitin monomers in total cell extracts due to a great abundance of poly and multi-ubiquitinated proteins. Recommended is size separation of protein extracts before gel electrophoresis focused on good resolution of region between 6-10 kDa.

Selected references Krasuska et al. (2016). Nitric oxide-polyamines cross-talk during dormancy release and germination of apple embryos. Nitric Oxide. 2016 Nov 23. pii: S1089-8603(16)30178-1. doi: 10.1016/j.niox.2016.11.003. [Epub ahead of print]

Application example

Western blot using anti-ubiqutin antibodies


The 10 μg of soluble protein extracts from 3 -week old Solanum tuberosum (1), one -week old seedlings of Lupinus angustifolius (2), Pisum sativum (3) , Zea mays L . (4) and Hordeum vulgare (5) leaves were extracted with buffer containing 50 м M Hepes- KOH, pH 7.5, 330 м M sorbitol, 2 м M EDTA, 1 м M MgCl 2, 5 м M ascorbate, 0.05% BSA . All samples were mixed with sample buffer, denatured for 5 min at 70°C and separated on 10% Tricine -SDS -PAGE . Proteins were blotted 30 min to HybondTM -PVDF membrane (Amersham Life Science) using semi -transfer (Bio -Rad) in standard transfer buffer in presence of 20% methanol. Transfer of proteins to the membrane was checked using 1% Ponceau S staining before the blocking step. Blots were blocked in buffer (5% low -fat milk in 1xPBS, 0. 1% Tween) with for 60 min at room temperature (RT) with agitation. Blots were incubated in the primary antibody at a dilution of 1: 5 000 at RT 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 P BS -T at RT with agitation. Blot was incubated in secondary antibody
(goat anti -rabbit IgG, AS09 602, Agrisera ) diluted to 1 :50 000 for 1h at RT with agitation. The blot was washed as above and developed for 5 min with Clarity Western ECL Substrate and ChemiDoc detection system (Bio-Rad).

Western blot using anti-ubiquitin antibodies

Total cell extracts of Solanum tuberosum (1), Lupinus angustifolius (2), Pisum sativum (3), Zea mays L. (4)Hordeum vulgare (5), were separated in conditions as described above. Marker - P restained SDS -PAGE ( Bio Rad, kat # 1610318). Monomers are marked by arrow. Exposure 30 seconds. For visualization of monomers, longer exposure time was applied.

Courtesy of Dr. Elena Poznidaeva Laboratory of Plant Gene Expression, Timiryazev Institute of Plant Physiology RAS, Moscow,  Russia


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