Rodius François, Hammer Claude, Vasseur Paule
Centre National de la Recherche Scientifique (CNRS) Formation de Recherche en Evolution (FRE) 2635 - ESE (Ecotoxicité, Santé Environnementale), Université de Metz, Metz, France.
Environ Mol Mutagen. 2004;43(4):209-16. doi: 10.1002/em.20021.
RNA-arbitrarily-primed PCR (RAP-PCR) can be used to detect changes in gene expression in organisms for which only minimal genomic information is available. In this study, RAP-PCR was used to detect modification of mRNA expression in the freshwater bivalve Unio tumidus, a mussel commonly used as a sentinel species in field studies. RNA expression was analyzed in the digestive glands of mussels from a control pond and in mussels transplanted into two sites on the Moselle River. The analysis identified a product in all animals exposed to sediments at the river station located downstream of a heavily populated area. This product was not present in animals exposed at the upstream station or at the control site. The additional PCR product was cloned and sequenced, and specific oligonucleotides for the sequence were designed to amplify cDNAs from control and transplanted mussels. A signal was obtained only with the cDNAs from animals exposed at the downstream station, confirming that the variation detected by RAP-PCR corresponds to an increase of gene expression. Chemical analysis of sediments from the control and river sites indicated that the levels of several potential pollutants were similar at the three locations and below currently accepted pollution thresholds. Our results indicate that RAP-PCR is a sensitive technique that can be applied in field studies to identify modifications in the gene expression of bioindicator species. This approach can complement chemical, biochemical and population studies to assess the impact of human activity on the ecological quality of aquatic systems.
RNA任意引物PCR(RAP-PCR)可用于检测那些仅有极少基因组信息的生物体中基因表达的变化。在本研究中,RAP-PCR被用于检测淡水双壳贝类膨背蚌(一种在野外研究中常用作指示物种的贻贝)中mRNA表达的变化。对来自对照池塘的贻贝以及移植到摩泽尔河两个地点的贻贝的消化腺中的RNA表达进行了分析。分析在所有暴露于人口密集区下游河段沉积物的动物中鉴定出一种产物。在暴露于上游河段或对照地点的动物中未发现该产物。将额外的PCR产物进行克隆和测序,并设计针对该序列的特异性寡核苷酸以扩增对照贻贝和移植贻贝的cDNA。仅从暴露于下游河段的动物的cDNA中获得了信号,证实RAP-PCR检测到的变化对应于基因表达的增加。对对照地点和河段沉积物的化学分析表明,三个地点几种潜在污染物的水平相似且低于目前公认的污染阈值。我们的结果表明,RAP-PCR是一种敏感技术,可应用于野外研究以鉴定生物指示物种基因表达的变化。这种方法可以补充化学、生化和种群研究,以评估人类活动对水生系统生态质量的影响。