AI Article Synopsis
- - Auxin measurements, particularly of indole-3-acetic acid (IAA), are essential for understanding plant signaling and metabolic balance.
- - This protocol utilizes gas chromatography-mass spectrometry (GC-MS) for high-throughput analysis of free IAA, allowing for the processing of over 500 samples weekly with minimal tissue required.
- - The procedure involves several steps including plant harvesting, purification, derivatization, and analysis, taking about 3 hours for the first eight samples, with time efficiency increasing for larger batches when using robotic methods.
Article Abstract
Auxin measurements in plants are critical to understanding both auxin signaling and metabolic homeostasis. The most abundant natural auxin is indole-3-acetic acid (IAA). This protocol is for the precise, high-throughput determination of free IAA in plant tissue by isotope dilution analysis using gas chromatography-mass spectrometry (GC-MS). The steps described are as follows: harvesting of plant material; amino and polymethylmethacrylate solid-phase purification followed by derivatization with diazomethane (either manual or robotic); GC-MS analysis; and data analysis. [¹³C₆]IAA is the standard used. The amount of tissue required is relatively small (25 mg of fresh weight) and one can process more than 500 samples per week using an automated system. To extract eight samples, this procedure takes ∼3 h, whether performed manually or robotically. For processing more than eight samples, robotic extraction becomes substantially more time efficient, saving at least 0.5 h per additional batch of eight samples.
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| http://dx.doi.org/10.1038/nprot.2010.118 | DOI Listing |
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