At3g61400 Antibody
Description
Functional Characterization of AT3G61400
The AT3G61400 gene product belongs to the 2-oxoglutarate-dependent dioxygenase superfamily, which is involved in:
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Oxidative reactions requiring Fe²⁺ and ascorbate as cofactors .
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Secondary metabolite biosynthesis, including pathways for flavonoids and phytohormones .
Transcriptomic studies in Arabidopsis autopolyploid lines revealed differential expression of AT3G61400 under stress conditions. For example:
| Condition | Expression Fold Change | Significance (p-value) |
|---|---|---|
| Wild-type (WT) | +1.67 | 0.0009 |
| rpt2 mutant | +1.40 | 0.0125 |
This upregulation suggests a role in stress adaptation, potentially through modulating oxidative pathways .
Transcriptome Profiling
At3g61400 Antibody has been employed in comparative transcriptome analyses to quantify protein levels in Arabidopsis mutants. Key findings include:
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Elevated expression in root tissues under phosphate starvation .
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Correlation with genes involved in lignin biosynthesis (e.g., 4CL4, PGL5) .
Protein Localization Studies
Immunohistochemistry (IHC) using this antibody localized the AT3G61400 protein to vascular tissues, supporting its hypothesized role in xylem differentiation .
Technical Considerations
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Specificity: The antibody’s epitope resides in the N-terminal region (residues 15-120) of the AT3G61400 protein .
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Validation: Western blot analyses confirm a single band at ~35 kDa, consistent with the predicted molecular weight .
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Limitations: Cross-reactivity with homologous dioxygenases in Brassica species has been observed, necessitating species-specific controls .
Future Directions
While current data emphasize AT3G61400’s metabolic roles, further studies could explore:
Product Specs
Constituents: 50% Glycerol, 0.01M Phosphate Buffered Saline (PBS), pH 7.4
Q&A
FAQs for Researchers on At3g61400 Antibody
Advanced Research Questions
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How to resolve contradictions in At3g61400 antibody cross-reactivity across plant species?
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Troubleshooting Framework:
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Hypothesis Testing: Cross-reactivity may arise from conserved epitopes in homologous proteins (e.g., Brassica species).
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Experimental Fix: Perform phylogenetic alignment of At3g61400 homologs to identify conserved regions and design peptide-blocking assays .
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Data Interpretation: Use competitive ELISA with peptides from non-target species to quantify cross-reactivity thresholds.
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What strategies address low antibody affinity in functional studies of At3g61400?
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Advanced Solutions:
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Epitope Mapping: Identify antibody-binding regions using peptide arrays or hydrogen-deuterium exchange mass spectrometry (HDX-MS) .
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Affinity Maturation: Apply phage display or yeast surface display to engineer higher-affinity variants .
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Negative Control: Include At3g61400 knockout lines in immunoprecipitation (IP) assays to rule off-target interactions .
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How to design a study investigating At3g61400’s role in stress responses?
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Multidisciplinary Workflow:
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Phenotypic Screening: Expose At3g61400 mutants to abiotic stressors (e.g., drought, salinity).
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Transcriptomic Profiling: Compare RNA-seq data between mutants and wild-type under stress.
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Antibody-Based Protein Quantification: Use quantitative Western blotting to correlate protein levels with stress phenotypes .
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Data Contradiction Analysis
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How to reconcile conflicting reports on At3g61400’s substrate specificity?
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Root-Cause Investigation:
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Variable Sources: Differences in recombinant protein purification methods (e.g., inclusion bodies vs. soluble expression).
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Mitigation: Standardize assays using uniformly purified protein and include positive controls (e.g., known 2OG oxygenase substrates).
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Evidence Synthesis:
Study Substrate Identified Method Potential Confounder A Flavonoids LC-MS Contaminating plant extracts B Alkaloids NMR Non-physiological pH conditions
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