BioTechniques News
Tristan Free
“It’s a bit like giving the soil a DNA test.” A new droplet digital PCR (ddPCR) method quantifies root DNA directly from soil samples, providing an invaluable insight into plant biomass and carbon storage.
Lurking beneath the ground, plant roots harbor an abundance of secrets, which a new ddPCR platform may help to uncover. Dubbed a ‘DNA test for soil’, the method was developed by researchers at Aarhus University (Denmark) and provides a way to substantially simplify and improve root species profiling and biomass quantification, with implications for plant breeding, as well as climate science and biodiversity research.
Roots are essential for plant survival and growth, and represent a major route for carbon sequestration, which is becoming increasingly important in the face of climate change. Unfortunately, due to their subterranean location, roots can be tricky to study. Traditional methods for root species profiling and biomass quantification, such as root separation and C13 labeling, are time- and labor-consuming and can be inaccurate, opening the door to DNA-based techniques.
In previous work, quantitative real-time PCR has been used to identify the species diversity of roots in mixed soil samples, although this approach is hindered by sensitivity to PCR inhibitors and primer efficiency variability. To take this field forward, the team behind the recent research turned to ddPCR, hoping to achieve precise and efficient quantification of root biomass that overcomes the limitations of existing methods.
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The resulting ddPCR assay targeted the genetic marker Internal Transcribed Spacer (ITS2) – a ubiquitous ribosomal DNA barcode. Universal primers were used to target this region in grasses, while modified primers were designed for legumes and forbs. DNA was extracted from soil samples, before being partitioned into nanoliter droplets using the QX200 Automated Droplet Generator from Bio-Rad (CA, USA). PCR was performed in a C1000 Touch Thermal Cycler, also from Bio-Rad, and included 10 minutes at 95 °C followed by 39 cycles, consisting of: 30 seconds denaturation at 94 °C, annealing for 1 minute at 55 °C and extension for 30 seconds at 72 °C.
After this amplification, the researchers were able to identify individual species from soil samples, as well as estimate total DNA content and, subsequently, root biomass.
“It’s a bit like giving the soil a DNA test,” explained study author Henrik Brinch-Pedersen. “We can suddenly see the hidden distribution of species and biomass without digging up the whole field.”
This ddPCR-based strategy has several advantages over other methods, including enhanced sensitivity and absolute quantification of target DNA, increased accuracy and reliability, and the ability to quantify roots directly from soil containing a mixture of different species.
The approach has a number of potential uses in plant and climate science, Brinch-Pedersen commented: “We see great potential in using this method to develop varieties that store more carbon in the soil. It could become an important tool in future agriculture.”
Next, the researchers hope to broaden the scope of their test, expanding their primer–probe sets to include additional plant species, so that in the future, they can measure even more species directly from soil samples.
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