GAI Antibody
Description
Definition and Biological Context
The GAI protein is a DELLA-domain transcriptional repressor in Arabidopsis thaliana that regulates GA-mediated processes such as seed germination, stem elongation, and flowering . The GAI Antibody specifically targets epitopes within this protein, enabling researchers to study its expression, localization, and interactions . DELLA proteins like GAI function as negative regulators of GA signaling, and their degradation via the 26S proteasome is central to GA response pathways .
Antibody Development and Design
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Immunogen: A synthetic peptide derived from the Arabidopsis GAI protein sequence (UniProt: Q9LQT8) .
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Reactivity: Specific to Arabidopsis thaliana; no cross-reactivity observed in mutant backgrounds (e.g., gai mutants) .
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Applications:
Protein Localization and Expression Analysis
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The GAI Antibody has been used to study subcellular localization in Arabidopsis roots, revealing cytoplasmic and nuclear distribution patterns .
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In GA-treated gid1 (GA receptor) mutants, the antibody demonstrated GAI protein stabilization, confirming its role in GA signaling .
Functional Studies in Mutant Backgrounds
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Western blot analyses using the GAI Antibody showed no detectable signal in gai mutants, validating its specificity .
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Immunoprecipitation studies coupled with mass spectrometry have explored GAI’s interaction partners, such as GA receptors (GID1) and ubiquitin ligases (SLY1) .
Technical Validation and Challenges
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Affinity Purification: Critical for improving detection rates. For example, recombinant protein-purified GAI Antibodies achieved a 55% success rate in detecting target proteins .
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Limitations: Low-abundance targets may require signal amplification, and nonspecific binding was observed in rare cases (e.g., faint signals in pin3 mutants) .
Table 2: Antibody Characterization Data
| Parameter | Detail | Source |
|---|---|---|
| Molecular Weight | 58.9 kDa (expected) | |
| Host Species | Rabbit | |
| Cross-Reactivity | None in gai mutants | |
| Storage Conditions | Lyophilized at -20°C |
Broader Implications in Antibody Reproducibility
The GAI Antibody’s development aligns with efforts to address the "antibody characterization crisis" in plant research . Initiatives like the Global Biological Standards Institute (GBSI) emphasize rigorous validation to ensure reproducibility, particularly for low-abundance proteins like GAI .
Future Directions
Product Specs
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4
Target Background
- GAI negatively regulates dark-induced senescence and chlorophyll degradation. PMID: 29574486
- The DELLA protein, GAI, undergoes mono-O-fucosylation by the novel O-fucosyltransferase SPINDLY (SPY) in Arabidopsis thaliana. This O-fucosylation activates DELLA by promoting its interaction with key regulators in brassinosteroid- and light-signaling pathways. PMID: 28244988
- This study establishes a direct link between melatonin and floral transition, indicating the novel involvement of DELLAs-activated FLC in melatonin-mediated flowering in Arabidopsis. PMID: 26887824
- BBX24 physically interacts with DELLA proteins and alleviates DELLA-mediated repression of PIF4 activity. PMID: 25656233
- Our data suggest that the miR171-SCL module is critical for mediating GA-DELLA signaling in the coordinate regulation of chlorophyll biosynthesis and leaf growth in light. PMID: 25101599
- Transcriptional analysis supported these findings and revealed that a complex signaling network has responded to MgT in Arabidopsis. DELLA enhancement, which depends on ABI1, contributed to the remodeling growth and development of young seedlings. PMID: 25074907
- This research determined the TF interactome of the Arabidopsis (Arabidopsis thaliana) DELLA protein GIBBERELLIN INSENSITIVE and screened a collection of conditional TF overexpressors in search of those that alter GA sensitivity. PMID: 25118255
- RGA and GAI play distinct roles in seed germination, and GAI interacts with DAG1 in embryo development. PMID: 25064446
- The study showed that the WD-repeat/bHLH/MYB complex acts as a direct target of DELLAs in the GA pathway and that both DELLAs and JAZs interacted with the WD-repeat/bHLH/MYB complex to mediate synergism between GA and JA signaling. PMID: 24659329
- The removal of DELLA repression highly upregulated certain genes on various hormone pathways, suggesting that gibberellin signaling acts upstream of the jasmonic acid, salicylic acid, and ethylene pathways in regulating leaf senescence. PMID: 24576761
- Further analyses demonstrated that TOPP4 is able to directly bind and dephosphorylate RGA and GAI, confirming that the TOPP4-controlled phosphorylation status of DELLAs is associated with their stability. PMID: 25010794
- The DELLA protein functions as a transcriptional activator through the DNA binding of the indeterminate domain family proteins. PMID: 24821766
- Atgai mRNA can move from companion cells to sieve tubes, and the translated product remains at the sites to which it is transported. PMID: 24144190
- GAI requires BOTRYTIS SUSCEPTIBLE1 INTERACTORs (BOIs) to inhibit GA responses such as seed germination, phase transition, and flowering. PMID: 23482857
- Results show that the induction of GAI activity has an almost immediate effect on gene expression. PMID: 21904598
- Through transgenic modification, DELLA-less versions of GAI (gai) and RGL1 (rgl1) in a Populus tree have profound, dominant effects on phenotype, producing pleiotropic changes in morphology and metabolic profiles. PMID: 16404575
- PIL5 binds directly to GAI promoters but not to Giberellic and Abscisic metabolic gene promoters in Arabidopsis. PMID: 17449805
- Data show that gai gene apple transgenic clones showed reduced rooting ability, especially for the extremely compact ones. PMID: 17932677
- A study evaluating ethylene-giberillic acid crosstalk in gibberellin-insensitive (gai), ethylene-overproducing 2-1 (eto2-1) double mutant A. thaliana is reported. PMID: 18078472
- The RNA mobile elements in GAI RNA play important roles in RNA movement. PMID: 19453448
