Highly Sensitive and High-Throughput Analysis of Plant Hormones Using MS-Probe Modification and Liquid ChromatographyTandem Mass Spectrometry: An Application for Hormone Profiling in Oryza sativa
Highly Sensitive and High-Throughput Analysis of Plant Hormones Using MS-Probe Modification and Liquid ChromatographyTandem Mass Spectrometry: An Application for Hormone Profiling in Oryza sativa
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DOI：
10.1093/pcp/pcp057
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发表时间：
2009-07-01
影响因子：
4.9
通讯作者：
Sakakibara, Hitoshi
中科院分区：
文献类型：
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作者：
Kojima, Mikiko;Kamada-Nobusada, Tomoe;Sakakibara, Hitoshi
我们建立了一种高灵敏度和高通量的方法，用于同时分析43种细胞分裂素、生长素、阿坝和赤霉素。该方法包括用于固相萃取和超高效液相色谱法（UPLC）的自动液体处理系统，以及配备电喷雾接口（ESI; UPLC-ESI-qMS/MS）的串联四极杆质谱仪（qMS/MS）。为了提高带负电荷的化合物，如赤霉素的检测限，我们化学衍生级分含有生长素，阿坝和赤霉素与溴胆碱，具有季铵官能团。我们称之为MS探针的这种修饰使这些激素衍生物具有正离子电荷，并允许在单次运行中使用UPLC-ESI-qMS/MS以正离子模式测量所有化合物。因此，赤霉素的定量限增加到50倍。我们目前的方法需要<100 mg（FW）的植物组织来确定植物激素谱，并使我们能够同时分析180个植物样品。将该方法应用于植物激素谱分析，使我们能够绘制水稻激素种类的器官分布图，并确定水稻赤霉素信号突变体gid 1 -3、gid 2 -1和slr 1中四种主要激素之间的相互作用。将激素谱数据的结果与赤霉素信号转导突变体中的转录组数据相结合，使我们能够分析基因表达变化与激素代谢之间的关系。
We have developed a highly sensitive and high-throughput method for the simultaneous analysis of 43 molecular species of cytokinins, auxins, ABA and gibberellins. This method consists of an automatic liquid handling system for solid phase extraction and ultra-performance liquid chromatography (UPLC) coupled with a tandem quadrupole mass spectrometer (qMS/MS) equipped with an electrospray interface (ESI; UPLC-ESI-qMS/MS). In order to improve the detection limit of negatively charged compounds, such as gibberellins, we chemically derivatized fractions containing auxin, ABA and gibberellins with bromocholine that has a quaternary ammonium functional group. This modification, that we call MS-probe, makes these hormone derivatives have a positive ion charge and permits all compounds to be measured in the positive ion mode with UPLC-ESI-qMS/MS in a single run. Consequently, quantification limits of gibberellins increased up to 50-fold. Our current method needs < 100mg (FW) of plant tissues to determine phytohormone profiles and enables us to analyze 180 plant samples simultaneously. Application of this method to plant hormone profiling enabled us to draw organ distribution maps of hormone species in rice and also to identify interactions among the four major hormones in the rice gibberellin signaling mutants, gid1-3, gid2-1 and slr1. Combining the results of hormone profiling data with transcriptome data in the gibberellin signaling mutants allows us to analyze relationships between changes in gene expression and hormone metabolism.