Both DNA and RNA were simultaneously extracted from soil samples (2 g each), using a lab-made protocol based on phenol-chloroform-isoamylalcohol extraction (Harkes et al., 2019 (link)). Quality and quantity of the obtained RNA and DNA was measured with a Nanodrop and Qubit. The nucleic acid eluate was stored at -80°C until further processing. For synthesis of cDNA from extracted RNA, the Maxima First Strand cDNA Synthesis Kit for RT-qPCR (Fermentas, Thermo Fisher Scientific Inc., USA) was used according to the manufacturer’s instructions. All individual DNA and cDNA samples were diluted to 1 ng/µl and 0.1 ng/µl, respectively, and used as template for PCR amplification.
To estimate the nematode density, a subsample of the nematode suspension (1/10 of each sample) was counted under a dissecting microscope. This was done in triplicate. Hereafter, nematode suspensions were concentrated and lysed according to Vervoort et al. (2012) (link). This resulted in 100 µl purified DNA, which served as a template for quantitative PCR (qPCR).
To estimate the nematode density, a subsample of the nematode suspension (1/10 of each sample) was counted under a dissecting microscope. This was done in triplicate. Hereafter, nematode suspensions were concentrated and lysed according to Vervoort et al. (2012) (link). This resulted in 100 µl purified DNA, which served as a template for quantitative PCR (qPCR).
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