Auxin Detection Service

What Are Auxins

Auxins are a class of endogenous plant hormones characterized by the presence of an unsaturated aromatic ring and an acetic acid side chain. Their chemical essence is represented by indole-3-acetic acid (IAA) and includes other growth regulators such as indole-3-propionic acid, indole-3-butyric acid, and naphthaleneacetic acid (NAA). They play pivotal roles in regulating stem growth rates, suppressing lateral bud formation, and promoting root development.

Chemical Structure and Composition of Auxins

Auxins are characterized by their distinct chemical structure, comprising an unsaturated aromatic ring and an acetic acid side chain. The prototypical auxin, indole-3-acetic acid (IAA), possesses a core indole ring structure appended with a carboxyl group at the third carbon atom, denoted as the acetic acid side chain (Figure 1). This structural arrangement allows auxins to engage in various physiological processes that are crucial for plant growth and development.

Figure 1: Chemical structure of indole-3-acetic acid (IAA)

In addition to IAA, other compounds share structural similarities with auxins and are collectively referred to as auxin-like or auxin-related compounds. These include indole-3-propionic acid, indole-3-butyric acid, and naphthaleneacetic acid (NAA). While IAA remains the quintessential representative of auxins, these related compounds can also exert auxin-like effects, contributing to the complexity of hormonal regulation in plants.

Physiological Roles of Auxins:

Auxins play pivotal roles in orchestrating diverse physiological processes within plants. Some of their key functions include:

Regulation of Stem Growth: Auxins are instrumental in controlling the rate and direction of stem elongation. They achieve this by promoting cell elongation in the stem's apical meristem, leading to upward growth.

Suppression of Lateral Bud Formation: Auxins inhibit the development of lateral buds, ensuring that the plant directs its energy and resources toward primary shoot growth.

Promotion of Root Development: Auxins stimulate root growth and branching. They influence cell division and elongation in the root meristem, facilitating the establishment of an extensive root system.

Auxin Signal Transduction

Currently, researchers have identified three auxin signal transduction pathways in plants: (1) The Aux/IAA-TIR1 nuclear signaling pathway, which mediates auxin's transcriptional regulatory mechanism. When plant cells sense changes in external auxin concentration, it leads to alterations in gene transcription levels. These genes can be categorized into three families: SAUR, GH3, and Aux/IAA. Among them, Aux/IAA plays a central role in auxin transcriptional regulation. This protein can form dimers with auxin response factors (ARF), acting as inhibitors of auxin-induced genes, thereby negatively regulating auxin levels. TIR1 interacts with the second structural domain of the Aux/IAA protein, releasing ARF from the Aux/IAA-ARF dimer and positively regulating auxin levels [4]. (2) The cell surface-initiated signaling pathway, mediating rapid auxin responses. In this pathway, external auxin treatment can induce the phosphorylation of H+-ATPases on the cell membrane, facilitated by auxin-responsive genes SAUR (Small auxin-up RNA). This activates H+ transport out of the cell, increasing the acidity of the extracellular environment and acidifying the cell wall, promoting cell growth. The auxin-binding protein 1 (ABP1), located in the endoplasmic reticulum, can transport to the cell membrane and bind to auxin, completing auxin signal transduction. (3) The SKP2A signaling pathway, mediating cell cycle regulation. Auxin can trigger the degradation of the SKP2A protein, which promotes the degradation of cell cycle transcription factors E2FC and DPB, thereby regulating the cell cycle progression. Therefore, external auxin can mediate cell cycle regulation through the SKP2A protein. The specific regulatory processes of this pathway are still under investigation.

Accurate Quantification of Auxins

Creative Proteomics employs high-performance liquid chromatography (HPLC) and liquid chromatography-mass spectrometry (LC-MS) methods to efficiently and accurately detect changes in auxin levels. In addition, we offer other plant hormone detection services, including a high-throughput detection method based on the LC-MS/MS platform that covers six major classes of plant hormones, including auxins, gibberellins, cytokinins, abscisic acid, jasmonic acid, and salicylic acid. Quantitative analysis using isotope dilution enables the determination of absolute plant hormone content. Through efficient sample pretreatment techniques for the enrichment and purification of plant hormones, combined with the outstanding separation capabilities of ultra-high-performance liquid chromatography (UPLC) and the high selectivity of tandem mass spectrometry (MS/MS), interference from complex sample matrices is effectively reduced. This enhances the sensitivity of plant hormone detection and ensures the reliability of analytical results.

Sample Requirements

Cells: After sampling, centrifuge to remove the culture medium and immediately store in liquid nitrogen, then store at -80°C.

Leaves, stems, flowers, etc.: After removal, immediately place in liquid nitrogen, then store at -80°C.

Fruit flesh, fruit peel, etc.: If separation conditions are not available in the field, transfer the samples to an indoor setting as soon as possible (within 30 minutes), separate the samples, and place them in liquid nitrogen, then store at -80°C.

Mixed Sampling: Each sample should be a mixture from at least 3 or more tissues/plants. For example, in the case of rice leaves, take one leaf from the same position with similar phenotypes at the same time, simultaneously collect from 6 plants, place them in a 10 ml centrifuge tube, and rapidly freeze in liquid nitrogen.

Advantages of Auxins Analysis

We provide a full-service solution including sample preprocessing, analysis, and bioinformatics analysis, with comprehensive technical expertise and extensive experience.

Sample preprocessing involves liquid-liquid extraction in batches, resulting in a short analysis cycle.

Quantitative analysis is performed using UPLC-MS/MS, ensuring scientific accuracy through rigorous quality control (QC) measures.

Metabolomics analysis and metabolite-agronomic trait associations using kernels of wheat (Triticum aestivum) recombinant inbred lines.

Journal: Plant J.

Published: 2020 —

Method: Comprehensive Metabolomics + QTL Reveals the Association Analysis between Wheat Recombinant Inbred Line Grain Metabolites and Traits

Understanding the genetic basis of phenotypic traits has been a crucial goal in plant science. Plant metabolomics is often considered a bridge between the genome and phenotype, and its integration with quantitative genetic analysis helps infer genetic associations between plant metabolism and phenotypic variation. In this study, the authors utilized the wheat lines KN9204 and J411, which exhibit significant differences in grain and spike traits, to extract a total of 1,260 metabolites from mature grains. These metabolites mainly included flavonoids, polyphenols, plant hormones and derivatives, organic acids, sugars, and more. Broad-sense heritability data for metabolic traits indicated that the diversity of metabolites was primarily influenced by genetic factors. Linkage analysis revealed that 1,005 metabolic trait loci (methylation quantitative trait loci, mQTLs) were unevenly distributed throughout the entire genome. By considering compound structures, known biosynthetic pathways, and wheat genome annotations, 24 candidate genes were identified that could regulate the expression levels of different metabolites.

Correlation analysis was performed by integrating methylation quantitative trait loci (mQTLs) and phenotype QTLs (quantitative trait loci) for agronomic traits, revealing genetic relationships between metabolites and agronomic traits. Notably, two metabolites involved in auxin synthesis, mr1346 (tryptophan) and wm0034 (4-indoleacetaldehyde), were significantly positively correlated with the number of seeds per spike (NGPS). Further use of wheat metabolomics data to predict wheat agronomic traits demonstrated strong predictive capabilities for NGPS and plant height. This study suggests the potential use of metabolite information to assist in wheat breeding.

Metabolite-Agronomic Trait Association Network and Candidate Genes Related to Spikelet Number
(a) the network depicting associations between metabolites and agronomic traits, (b) the correlation between two specific metabolites and spikelet number, and (c) the QTL localization curves for spikelet number (NGPS) on chromosome 4B, as well as the candidate genes mr1346 (tryptophan) and wm0034 (4-indolecarboxaldehyde).

Reference

1. Shi T, Zhu A, Jia J, et al, Liu W, Ren X, Sun D, Fernie AR, Cui F, Chen W. Metabolomics analysis and metabolite-agronomic trait associations using kernels of wheat (Triticum aestivum) recombinant inbred lines. Plant J. 2020 Jul;103(1):279-292.