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cFBPase | Cytosolic fructose-1,6-bisphosphatase (cytoplasm marker in photosynthetic tissues)

AS04 043  |  Clonality: Polyclonal  |  Host: Rabbit  |  Reactivity: A. thaliana, B.napus, M. atropurpureum, N. benthamiana, P. silvestris, O. sativa, P. hybrida cv. Mitchell, S. tuberosum, Z. mays |  cellular [compartment marker] of cytoplasm 

cFBPase | Cytosolic fructose-1,6-bisphosphatase (cytoplasm marker in photosynthetic tissues) in the group Antibodies for Plant/Algal  / Carbohydrates at Agrisera AB (Antibodies for research) (AS04 043)

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Product Information

Immunogen

Overexpressed cytosolic fructose 1,6 bisphosphatase (cFBPase) derived from the sequence from Arabidopsis thaliana cFBPase UniProt: Q9MA79, TAIR: AT1G43670

Host Rabbit
Clonality Polyclonal
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)
Recommended dilution 1 : 5 000 (WB)
Expected | apparent MW

45 | 37 kDa (Arabidopsis thaliana)

Reactivity

Confirmed reactivity Arabidopsis thaliana, Brassica napus, Macroptilium atropurpureum, Nicotiana benthamiana, Pinus silvestris, Oryza sativa, Petunia hybrida cv. Mitchell, Solanum tuberosum, Zea mays cellular compartment marker of cytoplasm in photosynthetic tissues
Predicted reactivity Capsella rubella, Pisum sativum, Ricinus communis, Glycine max, Phaseolus vulgaris, Sesamum indicum, Spinacia oleracea, Populus trichocarpa, Vitis vinifera
Species of your interest not listed? Contact us
Not reactive in

Chlamydomonas reinhardtii

Application examples

Application examples Application example


Western blot using anti-cFBPase antibodies

10 µg of Arabidopsis thaliana Col-0 WT chloroplast total protein (1), 10µg Col-0 WT chloroplast stroma protein (2), Col-0 WT total leaf sample 1:10 dilution (3), Col-0 WT total leaf sample 1:2 dilution (4), Col-0 WT total leaf sample undiluted (5), Col-0 WT total leaf sample undiluted (6) and recombinat plastidial FBPase 0.05 µg, expressed in E.coli with no cTP present in the sequence (7), extracted with 2x Laemmli buffer and denatured at 95°C for 5 min. were separated on 10% SDS-PAGE and blotted to Millipore Immobilon-P membrain (carried out at 100 V for 90 min at 4°C in blotting buffer (25 mM Tris-HCl, 192 mM glycine, 10 % [v/v] methanol). Blots were blocked with blocking solution ( TBST buffer (20 mM Tris-HCl pH 7.4, 150 mM NaCl, 0.05 % [v/v] Tween-20) supplemented with 5 % [w/v] milk powder) for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 5 000 for 15h (over night) at 4°C with agitation in TBST. The antibody solution was decanted and the blot was washed 6 times for 10 min in TBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated) diluted to 1:20 000 in for 1h at RT with agitation. The blot was washed as above and ChemiGlow West Chemiluminescence substrate was used for development according to the manufacturer’s instructions and imaged using the ChemiDoc imaging system (Biorad, Cressier, France). Exposure time was 15 seconds.

Courtesy of Zanella Martina, ETH Zürich, Switzerland



Western blot using anti-cFBPase antibodies on A.thaliana roots

2 µg of total protein from Arabidopsis thaliana roots crude extract (1), supernatant after 10 0000 g centrifugation (2), extracted with ice-cold extraction buffer [50 mM Tris-HCl pH 7.5, 0.33 M Sucrose, 5 mM EDTA, 1x proteinase inhibitor] and denatured gradually with [50 mM Tris (pH 6.8), 10% Glycerol, 2% SDS, 2.5 M Urea, 0.005% Bromophenol Blue] at first with 55°C for 15 min followed by 95°C for 5 min. Proteins were separated on 12 % SDS-PAGE. Proteins were blotted 1h to PVDF using semi-dry transfer. Blots were blocked with 5 % nonfat milk + 0.1 % BSA for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 35 000 for ON/4°C with agitation in TBS-T. 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 RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase) diluted to 1:1 000 in for 2h at RT with agitation. The blot was washed as above and developed for 5 min withchemiluminescent detection reagent. Exposure time was 20 min.

Courtesy of Dr. Joanna Jeleńska and Dequantarius Speed, University of Chicago, USA

Additional information

Additional information

Kinetic and allosteric properties of the plant cytosolic FBPase are remarkably similar to the mammalian and yeast FBPase, but differ greatly from those of the chloroplastic FBPase. The antibody could detect FBPase from the human COS-7 cell line transfected with FBP1 expressing vector.

This product can be sold containing ProClin if requested.

This antibody does not react with chloroplastic form of FBPase.

Will this antibody be good as a cytosolic (non-microsomal control) in Arabidopsis thaliana roots? Although it has never been tested there is every likelihood that cFBPase will be expressed at reasonable levels even in roots. Even though the biosynthetic flux through to Sucrose may not be high as in mesophyll cells, central metabolism will still be active in young roots and the Sucrose etc being supplied externally still needs to be utilised. 

Related products

Background

Background

Fructose-1,6 bisphosphatase (FBPase) (EC=3.1.311)  is one of the regulatory enzymes in the sucrose biosynthetic pathway. In non-photosynthetic tissues, it regulates the rate of gluconeogenesis. In photosynthetic tissues, two FBPase isozymes (chloroplastic and cytosolic) play key roles in carbon assimilation and metabolism. In photosynthetic tissues cFBPase (cytosolic fructose 1,6 bisphosphatase) converts triose phosphates from the chloroplast to sucrose during light hours. Alternative name: D-fructose-1,6-bisphosphate 1-phosphohydrolase

Product citations

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