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PPDK | Pyruvate orthophosphate dikinase

AS13 2647 | Clonality: Polyclonal | Host: Rabbit | Reactivity: Arabidopsis thaliana, Oryza sativa, Zea mays

PPDK | Pyruvate orthophosphate dikinase in the group Antibodies for Plant/Algal  / Developmental Biology / Signal transduction at Agrisera AB (Antibodies for research) (AS13 2647)

DATA SHEET IN PDF

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How to cite this product:
Product name, number (Agrisera, Sweden)

Data sheet Product citations Protocols Add review

Product Information

Immunogen

Purified recomibinant enzyme consisting of residues 72-947 of Zea mays, UniProt: P11155

Peptide used to elicit this antibody is conserved in both isoforms of PPDK in rice: PPDK1 and PPDK2.
Host Rabbit
Clonality Polyclonal
Purity Affinity purified serum in PBS, pH 7.4
Format Lyophilized
Quantity 20 µg
Reconstitution For reconstitution add 200 µl of sterile water in 40% glycerol to a final protein concentration of 100 ng/µl
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)
Recommended dilution 1 : 25 000 (WB)
Expected | apparent MW

102 | 95 kDa

Reactivity

Confirmed reactivity Arabidopsis thaliana, Oryza sativa, Zea mays
Predicted reactivity Hordeum vulagre
Species of your interest not listed? Contact us
Not reactive in Cucumis sativus

Application examples

Application examples

Application example

western blot using anti-PPDK antibodies

5 µg of total protein from samples such as Zea mays leaf, were extracted with Protein Extraction 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 1:50 000 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 5 minutes.

Additional information

PPDK levels inr C3 plants like Arabidopsis thaliana and Hordeum vulgare are very low and PPDK protein is very dilute in most tissues of C3 plants. To perform detection in C3 plants leaf proteins needs to be concentrated before western blot, Chastain et al. (2002).

Related products

Background

Background
PPDK (Pyruvate, phosphate dikinase 1) is involved in formation of phosphoenolpyruvate and activated by light-induced dephosphorylation. Inhibited by dark-induced phosphorylation. PPDK is a low-abundance enzyme in C3 plants while it is a key enzyme of C4 photosynthesis.

Product citations

Selected references Shen et al. (2016). The existence of C4-bundle-sheath-like photosynthesis in the mid-vein of C3 rice. Rice (N Y). 2016 Dec;9(1):20. doi: 10.1186/s12284-016-0094-5. Epub 2016 May 10.

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