GOGAT | Glutamine oxoglutarate aminotransferase
AS07 242 | clonality: polyclonal | host: rabbit | reactivity: A.thaliana, N.tabacum, S. oleracea , O. sativa, Z. mays ,G. max , S. elongatus | plastidial marker
|Info:||Product suggestions||Add review|
1 : 1000 with standard ECL (WB)
|Expected | apparent MW||
177 | 170-180 kDa depending upon the species
Arabidopsis thaliana, Miscanthus giganteus, Nicotiana tabacum, Oryza sativa, Phaseolus vulgaris, Spartina sp., Zea mays
Arthrospira platensis PCC 7345, Beta vulgaris, Brachypodium distachyon, Burkholderia glumae, Camellia sinensis, Chloroflexi bacterium, Chlamydonas reinhardtii, Gracilaria tenustipitata, cyanobacteria, Crocosphaera watsonii WH 8502, Cyanidioschyzon merolae (strain 10D), Galdieria sulphuraria , Glycine max, Helicosporidium sp. ATCC 50920, Hydrogenobaculum sp. HO, Lactococcus lactis subsp. lactis bv. diacetylactis str. TIFN2, Leptospira interrogans serovar Bataviae str. HAI135, Medicago trucutula, Microcystis panniformis, Monoraphidium neglectum, Morus notabilis , Nicotiana tabacum, Ostreococcus lucimarinus, Leptolyngbya boryana, Physcomitrella patens subsp. patens , Populus trichocarpa, Porphyra purpurea, Sutterella wadsworthensis, Sulfurihydrogenibium yellowstonense, Veillonella atypica
|Not reactive in||
no confirmed exceptions from predicted reactivity known in the moment
A 40 kDa band present in A. thaliana sample is not competed away during antibody neutralization assay. In this assay free peptide used for antibody production is incubated together with anti-GOGAT antibodies. Due to the MW of this protein we suggest to use a gradient gel for protein separation and a longer transfer time.
|Selected references||Yang et al. (2016). Rice Ferredoxin-Dependent Glutamate Synthase Regulates Nitrogen-Carbon Metabolomes and Is Genetically Differentiated between japonica and indica Subspecies. Mol Plant. 2016 Sep 24. pii: S1674-2052(16)30195-2. doi: 10.1016/j.molp.2016.09.004.
Moscatelli et al. (2015). Characterisation of detergent-insoluble membranes in pollen tubes of Nicotiana tabacum (L.). Biol Open. 2015 Feb 20. pii: BIO201410249. doi: 10.1242/bio.201410249.
Podgórska et al. (2013). Long-term ammonium nutrition of Arabidopsis increases the extrachloroplastic NAD(P)H/NAD(P)+ ratio and mitochondrial reactive oxygen species level in leaves but does not impair photosynthetic capacity. Plant Cell Environ. April 10.
20 µg of total protein from (1) Spartina patens total cell extracted with Protein Extration Buffer, PEB (AS08 300), (2) Spartina alterniflora total cell, extracted with PEB, (3) Miscanthus giganteus total cell extracted with PEB, (4) Zea mays total cell extracted with PEB, (5) Phaseolus vulgaris total cell extracted with PEB, (6) Arabidopsis thaliana 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: 10 000 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, from Abcam) 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).
Approximately 40 µg of total protein from Arabidopsis thaliana wild-type (WT) and gls1-103 mutant (M) leaves, the latter of which showed reduced GOGAT levels, was extracted with a protein extraction solution containing 50mM Imidazole/HCl (pH 7.0 at 4ºC), 20% glycerol, 5mM 6-aminocaproic acid, 1mM EDTA. Protein extracts were then separated on 6% SDS-PAGE and blotted overnight at 4ºC to PVDF using the tank transfer method. Blots were blocked with PBS-T containing 5% skim milk for 1h at room temperature (RT) with agitation. Blot was incubated in the primary antibody at a dilution of 1: 20,000 in PBS-T containing 1% (w/v) BSA for 1h 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 PBS-T at RT with agitation. Blot was incubated in secondary antibody (anti-rabbit IgG horse radish peroxidase conjugated, from Sigma A9169) diluted to 1:20 000 in PBS-T containing 1% (w/v) BSA for 1h at RT with agitation. The blot was washed as above and developed for 1 min with Western Lightening Plus (Perkin-Elmer) according to the manufacturer's instructions. Exposure time was 5 min.
Courtesy Dr. Atsushi Takabayashi, Institute of Low Temperature Science, Hokkaido University, Japan
||| For other applications or usage on species other than stated above Agrisera offers a payment-after-testing option. To learn more about this or for any questions please use the LiveChat option or contact us at firstname.lastname@example.org