浮游动物DNA宏条形码标志基因比较研究

DNA宏条形码技术作为一种新型生物监测方法,在未来生态环境监测中有巨大的应用潜力。目前,浮游动物DNA宏条形码监测仍在发展阶段,需要首先对其(采样方法、引物选择和数据分析等)进行标准化和调整,然后才能用于常规流域生态监测。其中,如何选择合适的PCR扩增引物是DNA宏条形码生物监测标准化的关键问题之一。本研究比较了COI、18SV9和16S通用引物在浮游动物DNA宏条形码监测中的差异,为初步建立规范化的浮游动物DNA宏条形码监测方法提供技术支撑。结果表明,16S引物对浮游动物具有更好的特异性,其产生的操作分类单元(operational taxonomic unit, OTU)有88.1%属于浮游动物。虽然18SV9引物具有更高的物种覆盖度,不仅能扩增出浮游动物,还能扩增出大量藻类和真菌,但其物种识别敏感性较差,不适合浮游动物物种水平多样性监测。COI引物的浮游动物物种特异性、物种覆盖度和物种识别敏感性都适中,检出的浮游动物物种数量高于18SV9引物和16S引物,更加适合浮游动物DNA宏条形码监测。

Study on the Selection of Marker Genes in Zooplankton DNA Metabarcoding Monitoring

As a new biomonitoring method, environmental DNA (eDNA) metabarcoding has great potential to be applied in the future environmental monitoring of aquatic ecosystem. To promote its application in the routine biomonitoring, it is urgent to establish standardized methods and technical protocols for eDNA metabarcoding. Selection of suitable PCR primers is one of the key issues in eDNA based biomonitoring. This study evaluated the differences of PCR primers (18SV9, COI, 16S) in zooplankton biodiversity profiling by eDNA metabarcoding, and provided technical details for the establishment of standardized zooplankton monitoring approach. The results showed that the 16S primer had better specificity to zooplankton species, where 88.1% of 16S OTU (operational taxonomic unit) came from zooplankton. Although 18SV9 primer had higher species coverage than COI and 16S (besides zooplankton, a number of algae and fungi species also found), but the species identification sensitivity was lower. COI primers found 92 different zooplankton species, which was the largest number among the 3 pairs of primers. Overall, COI primer is more suitable for the study of zooplankton monitoring due to its moderate specificity to zooplankton species, species coverage and species identification sensitivity.