A PCR-based method for the identification of the roots of 10 co-occurring grassland species in mesocosm experiments

Abstract

An understanding of factors influencing the distribution of plant roots is intimately linked to our understanding of basic ecosystem functions such as nutrient flux and productivity. However, it is not usually possible to measure root distributions because it is difficult to identify the roots of different species when they are grown in mixture. This is because the roots of most species are not visually distinguishable. We designed a simple, PCR-based method for the identification of roots in mesocosm experiments, which we have applied to 10 co-occurring grassland species. Species-specific primers based on ITS sequences from GenBank were evaluated in PCR assays using either homogeneous or heterogeneous DNA templates, as well as DNA extracted from mixed-root samples from multiple combinations of species. The species-specific primers reported here produced accurate identifications, free from both false negatives and false positives, in 100% of our assays. We also evaluated the sensitivity of our system and demonstrated detection of species when they comprised as little as 0.05 ng of target DNA mixed in a total of 2.5 ng of multi-species template DNA. Our PCR-based method for root identification in mesocosms is more cost effective, and simpler to apply than previously described methods.