Molecular diagnostic assays have been developed and utilized to diagnose and to confirm the diagnoses of many plant-parasitic nematodes. We screened several gene sequences of soybean cyst nematode (SCN) [Heterodera glycines, Ichinohen] for their use as molecular markers. A methodology then was developed to use them to detect and quantify the number of H. glycines directly from Mississippi soil. A novel procedure utilizing multiple databases containing nematode DNA and EST sequences was developed to assist in the selection of SCN primers used in the PCR and qPCR assays. In vitro testing demonstrated that the DNA primers and probes developed from the novel procedure for the qPCR assays could accurately detect the presence of SCN. Subsequent testing resulted in a trend of increasing observed numbers of SCN contributing to increasing estimates by qPCR.