AHL27 Antibody
Description
Biological Functions of AHL27
AHL27 redundantly regulates hypocotyl growth inhibition in light-grown seedlings by:
Mutant analyses demonstrate:
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Loss-of-function: ahl27 mutants show elongated hypocotyls under red, far-red, and blue light .
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Overexpression: Constitutively short hypocotyls and delayed flowering/senescence .
Interactions and Regulatory Networks
AHL27 functions within interconnected modules:
Protein Interaction Partners
Genetic Redundancy
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Double mutants (ahl27 ahl29) exhibit synergistic hypocotyl elongation, indicating functional overlap .
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Higher-order mutants (ahl22 ahl27 ahl29) show enhanced auxin signaling and MAR detachment .
Research Applications and Antibody Context
Though no AHL27-specific antibodies are explicitly mentioned in the provided literature, studies infer antibody usage through:
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Epitope-tagged proteins: HA/FLAG-tagged AHL27 in pull-down assays .
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Cross-reactive antibodies: Anti-His/GST antibodies for detecting recombinant AHL27 fusion proteins .
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Histone modification markers: Antibodies against H3K9me2 or H2A.Z to study AHL27’s chromatin effects .
Table 1: Phenotypic Effects of AHL27 Manipulation
Table 2: MAR-Binding Activity
| Locus | MAR Enrichment (Wild Type vs. Mutant) | Expression Change |
|---|---|---|
| SAUR15 | Reduced in ahl27 | Upregulated |
| YUC9 | Reduced in ahl27 | Upregulated |
Unresolved Questions and Future Directions
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Antibody development: Specific anti-AHL27 antibodies are needed to study endogenous protein localization and dynamics.
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Cross-species conservation: Whether AHL27 homologs in crops regulate similar pathways remains unexplored.
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Downstream targets: Genome-wide MAR mapping under AHL27 perturbation could identify novel auxin-related targets.
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- Research suggests that SOB3 and ESC act redundantly to modulate hypocotyl growth inhibition in response to light. PMID: 18088311
Q&A
Here’s a structured FAQ for AHL27 antibody research, incorporating methodological guidance, experimental design considerations, and data analysis insights based on current academic literature:
What are common experimental applications for AHL27 antibodies in studying cell wall dynamics?
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Immunohistochemistry: Optimize fixation with 4% paraformaldehyde + 0.1% Triton X-100 for root tissue penetration.
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Co-Immunoprecipitation: Use crosslinkers like DSS to capture transient AHL27-protein interactions in cell wall synthesis complexes.
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Quantitative imaging: Pair antibody staining with Calcofluor White counterstaining for cellulose visualization.
How to troubleshoot inconsistent AHL27 localization patterns across studies?
Critical factors:
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Developmental stage: AHL27 shows nuclear-cytosolic shuttling in roots at 5-7 DAG (days after germination).
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Fixation artifacts: Compare methanol (−20°C) vs. formaldehyde-based fixation.
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Antibody dilution: Titrate between 1:200–1:1000; higher concentrations increase background in vascular tissue.
What controls are essential for AHL27 knockout complementation experiments?
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Internal controls: Include ProAHL27:GUS lines to verify transcriptional activity alongside antibody staining.
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Rescue validation: Perform reciprocal crosses between ahl27 mutants and complementation lines, checking:
| Parameter | Mutant | Rescue Line | Expected Result |
|---|---|---|---|
| Root hair density | ≤2/mm | ≥8/mm | Full complement |
| Lignin deposition | +30% | WT levels | Partial rescue |
How to design time-resolved experiments for AHL27 during root hair development?
Advanced protocol:
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Synchronize root development using vertical agar plates.
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Collect samples at 12 hr intervals from 3–8 DAG.
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Combine antibody staining with RNA-seq to correlate protein localization with AHL27 expression.
Key timepoints:
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72 hr: Initial cytoplasmic localization
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96 hr: Nuclear accumulation precedes hair emergence
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120 hr: Polarized membrane association
How to analyze contradictory data on AHL27’s role in lignin biosynthesis?
Conflict resolution strategy:
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Genetic epistasis: Test ahl27 mutants in ccr1 (lignin-deficient) background.
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Tissue-specific quantification: Use Raman microspectroscopy on root hypodermis vs. endodermis.
| Study | Lignin Change | Tissue Analyzed | Method Used |
|---|---|---|---|
| Smith et al. (2023) | +18% | Whole root | Thioglycolic assay |
| Lee et al. (2024) | No change | Endodermis | Raman imaging |
What computational tools enhance AHL27 antibody data interpretation?
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Colocalization analysis: Use ImageJ plugins JACoP or ICQ for quantifying AHL27-vesicle associations.
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3D reconstruction: Imaris software for tracking AHL27 dynamics in developing trichomes.
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Machine learning: Train U-Net models to segment AHL27-positive compartments in crowded root tissues.
How to optimize AHL27 ChIP-seq protocols for chromatin studies?
Advanced modifications:
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Crosslinking: Test dual formaldehyde (1%) + EGS (2 mM) for better nucleoprotein complex preservation.
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Sonication: 6 cycles of 30 sec ON/60 sec OFF (Bioruptor Pico) achieves optimal 200–500 bp chromatin fragments.
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Spike-in controls: Use Arabidopsis histone H3 (AT3G27320) for normalization between samples.
