Customize FAX1 Antibody

Description

Definition and Purpose of FAX1 Antibody

The FAX1 antibody is a polyclonal or monoclonal antibody designed to detect and quantify FAX1 protein levels in plant tissues. It is primarily used to:

Study FAX1’s role in plastid fatty acid export and lipid metabolism
Validate genetic mutants (e.g., fax1 knockouts or overexpressors)
Investigate FAX1 localization and interaction partners

3.1. Mutant Validation

Immunoblot analyses using FAX1 antibody confirmed reduced FAX1 levels in fax1 Arabidopsis mutants and overexpression in transgenic lines .
Example: fax1 knockouts showed a 50–70% reduction in C29 ketone waxes in stem cuticles .

3.2. Subcellular Localization

FAX1 antibody enabled precise localization studies, confirming its presence in chloroplast inner envelopes but not in ER membranes .
In Chlamydomonas, Cr-FAX1 localized to chloroplast membranes, while Cr-FAX5 (a homolog) associated with microsomes .

3.3. Stress Response Studies

FAX1 degradation under cold stress was tracked using the antibody, revealing a 50% reduction in wild-type Arabidopsis after 7 days at 4°C .
RBL11 protease mutants (rbl11) showed elevated FAX1 levels, linking cold sensitivity to FAX1 accumulation .

4.1. Role in Lipid Metabolism

FAX1 knockout lines exhibited:
- Reduced ER-derived lipids (e.g., phosphatidylcholine)
- Accumulation of plastid lipids (e.g., monogalactosyldiacylglycerol)
Overexpression increased triacylglycerol (TAG) by 30–40% in leaves and flowers .

4.2. Pollen Development

FAX1 antibody revealed defective pollen wall structures in fax1 mutants due to impaired sporopollenin and tryphine deposition .

4.3. Interaction with Proteases

FAX1 degradation by rhomboid protease RBL11 under cold stress was quantified, showing:

Genotype FAX1 Level at 4°C
Wild-type 20% of initial
rbl11 80% of initial

Genotype	FAX1 Level at 4°C
Wild-type	20% of initial
rbl11	80% of initial

Challenges and Limitations

Cross-Reactivity: FAX1 antibody may detect other FAX isoforms (e.g., FAX2–FAX4) due to sequence homology .
Quantitative Variability: Differences in protein extraction efficiency from chloroplast envelopes can affect immunoblot accuracy .

Product Specs

Buffer

Preservative: 0.03% Proclin 300
Constituents: 50% Glycerol, 0.01M PBS, pH 7.4

Form

Liquid

Lead Time

Made-to-order (14-16 weeks)

Synonyms

FAX1 antibody; At3g57280 antibody; F28O9.130Protein FATTY ACID EXPORT 1 antibody; chloroplastic antibody; At-FAX1 antibody

Target Names

FAX1

Uniprot No.

Q93V66

Target Background

Function

FAX1 Antibody mediates the export of free fatty acids from plastids. It exhibits a preference for palmitic acid (C16:0) over oleic acid (C18:1) and stearic acid (C18:0). It is not involved in fatty acid activation. FAX1 Antibody is essential for the biogenesis of the outer pollen cell wall, particularly for the assembly of exine and pollen coat, and for the release of ketone wax components.

Gene References Into Functions

Seed-specific overexpression of AtFAX1 has been shown to promote oil accumulation in Arabidopsis seeds. [AtFAX1] PMID: 29654768
FAX1 mediates fatty acid export from plastids. [FAX1] PMID: 25646734

Database Links

KEGG: ath:AT3G57280

STRING: 3702.AT3G57280.1

UniGene: At.909

Protein Families

TMEM14 family

Subcellular Location

Plastid, chloroplast inner membrane; Multi-pass membrane protein.

Tissue Specificity

Expressed in cotyledons, leaves, sepals and pollen.

Q&A

FAX1 Antibody Research FAQs for Academic Investigators

Advanced Research Questions

How to resolve contradictions in FAX1-mediated fatty acid transport across experimental systems?
- Methodological answer:
  - Yeast vs. plant systems: In yeast complementation assays, test FAX1’s ability to restore α-linolenic acid uptake in fat1 mutants under controlled conditions (e.g., 3.6 mM α-linolenic acid, 29-hour growth assays) . Note that FAX1 restores transport but not activation (unlike Faa1/Faa4) .
  - Cross-species validation: Express FAX1 orthologs (e.g., Brassica napus, Chlamydomonas) in Arabidopsis mutants to assess functional conservation .
  - Structural analysis: Use homology modeling of FAX1’s Tmemb_14 domain to predict membrane interaction sites, validated via site-directed mutagenesis .
What integrated omics strategies elucidate FAX1’s role in plant-microbe interactions?
- Methodological answer:
  - Multi-omics pipeline:
    1. Transcriptomics: Identify FAX1 co-expressed genes (e.g., lipid transporters, β-ketoacyl-CoA synthases) in public databases (e.g., Araport).
    2. Metabolomics: Quantify jasmonates or other FA-derived signaling molecules in fax1 root exudates.
    3. Microbiome profiling: Use 16S/ITS sequencing to compare microbial communities in fax1 vs. WT rhizospheres.
  - Experimental design: Grow plants under sterile vs. soil conditions to isolate FAX1-dependent lipid effects .

Technical Optimization Questions

How to troubleshoot FAX1 antibody cross-reactivity in non-model plant species?
- Methodological answer:
  - Epitope mapping: Compare FAX1 peptide sequences across species (e.g., residues 50–70 in Arabidopsis) to assess antibody compatibility.
  - Pre-absorption control: Incubate antibody with recombinant FAX1 protein prior to immunolocalization to confirm signal loss.
  - Alternative detection: Validate via CRISPR-Cas9-generated tagged lines (e.g., FAX1-HA) in target species .
What controls are essential for FAX1 functional studies in heterologous systems?
- Methodological answer:
  - Yeast assays: Include empty vector controls and faa1/faa4 double mutants to distinguish transport vs. activation .
  - Lipid trafficking assays: Use radiolabeled FAs (e.g., 14C-palmitate) with/without FAX1 expression, followed by TLC quantification .
  - Membrane topology: Perform protease protection assays on isolated chloroplasts to confirm FAX1’s inner envelope orientation .

Data Interpretation Guidelines

Conflicting localization data: If FAX1 signals appear in unexpected compartments (e.g., ER), re-validate using:
- Subcellular markers (e.g., TOC75 for chloroplasts, BiP for ER) .
- Fractionation + immunoblotting (e.g., microsomal vs. chloroplast proteins) .
Negative results in overexpression lines: Check for compensatory mechanisms (e.g., upregulated FAX2/3) via qRT-PCR .

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FAX1 Antibody