Spore trap system currently in use and that will be used in the proposed research to collect airborne spores of the downy mildew pathogens, Peronospora effusa and B. lactucae. A) The complete spore trap assembly with power box. Power is maintained by a 12 V battery within the box and a solar panel. B) Enlarged view of the spore trap head showing the removable stainless steel sampling rods (each 40 millimeters in length) connected to the rotating arm. The sampling rods are coated with grease for spore adherence.
By starting the PCR with known quantities of DNA along with unknown samples we can use a linear equation to interpolate the starting quantity of DNA in the unknown sample. In this case, our unknown samples are those collected from airborne sampling, spinach seed lots, or soil samples. The fewer PCR cycles it takes to reach a specific fluorescence level, the more DNA is present in the sample. The values of known seed or soil samples that are downy mildew-free will be spiked with P. effusa at various concentrations and compared to the values obtained by real-time PCR quantification of the pathogen in naturally infected/infested samples. This serves to calibrate the real-time PCR assay for pathogen quantification in the presence of PCR inhibitors that may be found in naturally infested samples. The DNA probe technology for DNA quantification follows similar principles, but in this case relies upon the quantification of fluorescence of a fluorophore-labeled probe that binds to the specific target DNA sequence.