浮游藻类环境DNA宏条形码监测引物的比较与验证

Comprehensive Comparison and Validation of eDNA Metabarcoding Primers for Phytoplankton Biomonitoring

环境DNA(eDNA)宏条形码技术通量高、重复性好,在未来生态环境监测中有巨大的应用潜力。目前,浮游藻类环境DNA监测仍处在发展阶段,尚缺乏统一的浮游藻类扩增引物。利用同一个野外环境样本,比较8对通用引物在浮游藻类环境DNA监测中的差异,为初步建立规范化的浮游藻类环境DNA监测方法提供支撑。结果表明,不同引物对浮游藻类扩增存在明显偏好性,靶向扩增16S rDNA的引物主要检出硅藻,其次是隐藻和绿藻;靶向扩增18S rDNA的1391、AD3和ANF 3对引物具有较高的浮游藻类扩增效率和物种辨识度,分别检出67、62、63个浮游藻属,其检出的浮游藻类的相对丰度排序均为硅藻>绿藻>隐藻>金藻>甲藻,可以作为通用引物用于浮游藻类环境DNA宏条形码监测。

Environmental DNA(eDNA)metabarcoding technology has high throughput and good repeatability,and has great application potential in the future ecological environment monitoring.At present,eDNA monitoring for phytoplankton is still in the development stage,and there is a lack of unified amplification primers.The differences of 8 pairs of universal primers in phytoplankton eDNA monitoring were comprehensive evaluated and compared with the same field environmental samples,which provided support for the initial establishment of standardized phytoplankton eDNA monitoring methods.The results showed that different primers had obvious preference for phytoplankton amplification.Primers targeting amplification of 16S rDNA mainly detected out Bacillariophyta,followed by Cryptophyta and Chlorophyta.The three pairs of primers 1391,AD3 and ANF,which targeted amplification of 18S rDNA,showed high amplification efficiency and species identification,and 67,62 and 63 genera of phytoplankton were detected,respectively.According to the relative abundances of sequences,ranking of the top five phytoplankton was Bacillariophyta>Chlorophyta>Cryptophyta>Chrysophyta>Dinophyta.Moreover,these three pairs of primers could be used as universal primers for phytoplankton eDNA metabarcoding monitoring.