背景:评估植物挥发性有机化合物(VOC)的种内变异涉及可能会偏向生物学解释的陷阱,特别是当几个实验室在联合项目上合作时。比较,实验室间环形试验可以说明此类分析的可重复性.
目的:在一项涉及五个实验室的环形试验中,我们研究了用聚二甲基硅氧烷(PDMS)收集VOC的可重复性,并通过热解吸-气相色谱-质谱(TD-GC-MS)进行了分析。作为模型植物,我们使用了Tanacetumvulgare,显示萜类化合物的显著多样性,形成所谓的化学型。我们用两种化学型进行了环形试验,以检查分析前植物VOC测量中技术变异的来源,分析,和分析后的步骤。
方法:在一个实验室中产生单克隆根插条,并分配给五个实验室,其中植物在实验室特定条件下生长。在茉莉酸(JA)处理之前和之后,在来自所有植物的PDMS管上收集VOC。此后,每个实验室(捐赠者)向其他四个实验室(接受者)发送了一组试管,用自己建立的程序进行TD-GC-MS。
结果:检测到VOC谱中的化学型特异性差异,但在供体和受体实验室中的总体差异很大。JA诱导的VOC谱变化是不可重复的。实验室特定的生长条件导致表型变异,从而影响所得的VOC谱。
结论:我们的环形试验表明,尽管努力使每个VOC测量步骤标准化,结果在质量和数量上都不同。我们的结果揭示了植物VOC研究中的变异来源,并可能有助于避免类似实验中的系统误差。
BACKGROUND: Assessing intraspecific variation in plant volatile organic compounds (VOCs) involves pitfalls that may bias biological interpretation, particularly when several laboratories collaborate on joint projects. Comparative, inter-laboratory ring trials can inform on the reproducibility of such analyses.
OBJECTIVE: In a ring trial involving five laboratories, we investigated the reproducibility of VOC collections with polydimethylsiloxane (PDMS) and analyses by thermal desorption-gas chromatography-mass spectrometry (TD-GC-MS). As model plant we used Tanacetum vulgare, which shows a remarkable diversity in terpenoids, forming so-called chemotypes. We performed our ring-trial with two chemotypes to examine the sources of technical variation in plant VOC measurements during pre-analytical, analytical, and post-analytical steps.
METHODS: Monoclonal root cuttings were generated in one laboratory and distributed to five laboratories, in which plants were grown under laboratory-specific conditions. VOCs were collected on PDMS tubes from all plants before and after a jasmonic acid (JA) treatment. Thereafter, each laboratory (donors) sent a subset of tubes to four of the other laboratories (recipients), which performed TD-GC-MS with their own established procedures.
RESULTS: Chemotype-specific differences in VOC profiles were detected but with an overall high variation both across donor and recipient laboratories. JA-induced changes in VOC profiles were not reproducible. Laboratory-specific growth conditions led to phenotypic variation that affected the resulting VOC profiles.
CONCLUSIONS: Our ring trial shows that despite large efforts to standardise each VOC measurement step, the outcomes differed both qualitatively and quantitatively. Our results reveal sources of variation in plant VOC research and may help to avoid systematic errors in similar experiments.