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Pipette 23.6.13 for ios download free8/23/2023 ![]() Because IAA is a mobile signaling molecule that can be transported among cells to form auxin gradients and auxin maxima that are essential for plant development, a method that allows quick and precise measurement of IAA in specific plant tissues will greatly facilitate understanding of auxin-regulated plant growth and development.Īmong the different pathways of IAA regulation, the biosynthesis of IAA is a central way to regulate cellular IAA levels and has been actively studied for the past six decades. Based on this functional mechanism, auxin responses in cells can be partially controlled by the cellular concentration of IAA. To trigger downstream signaling responses, IAA functions like molecular glue, which ties its receptor TIR1, an F-box protein, with Aux/IAA transcriptional repressors, leading to degradation of Aux/IAA and thus releasing the transcriptional suppression of auxin responsive genes. As the major form of natural auxin, indole-3-acetic acid (IAA) has been extensively studied, and mechanisms of its function and regulation are being revealed. The method will potentially advance knowledge of the role and regulation of IAA.Īuxin, the first discovered plant hormone, plays critical roles in plant growth, organ formation, and plant responses to environmental stimuli. The protocol can be used for measurement of these endogenous compounds using isotope dilution, and it can also be applied to analyze IAA biosynthesis and biosynthetic pathways using stable isotope labeling. The SPE tips and GC-MS/MS based method enables high-throughput and accurate quantification of IAA and its biosynthetic precursors from minute plant tissue samples. The amount of plant tissue required for the assay is small (typically 2–10 mg fresh weight), and the use of SPE tips is simple and convenient, which allows preparation of large sets of samples within reasonable time periods. The derivatized analytes are finally analyzed by selected reaction monitoring on a gas chromatograph-mass spectrometer (GC-MS/MS) to determine the precise abundance of analytes. The protocol starts with homogenizing plant tissues with stable-labeled internal standards added, followed by analyte purification using solid phase extraction (SPE) tips and analyte derivatization. We have developed high-throughput methods to quantify plant endogenous IAA and its biosynthetic precursors including indole, tryptophan, indole-3-pyruvic acid (IPyA), and indole-3-butyric acid (IBA). ![]() Meanwhile, pathways and genes involved in IAA biosynthesis are not fully understood, so it is necessary to analyze the production of IAA at the metabolite level for unbiased studies of IAA biosynthesis. Therefore, the precise quantification of local IAA concentration provides insights into the regulation of IAA and its biological roles. The signaling response to IAA is largely dependent on the local concentration of IAA, and this concentration is regulated by multiple mechanisms in plants. The plant hormone auxin, indole-3-acetic acid (IAA), plays important roles in plant growth and development.
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