This protocol describes the extraction of DNA from relatively large volumes of soil and subsequent real-time quantitative PCR (qPCR). The qPCR protocol is a modification of the protocol presented by : Wei, F., Fan, R., Dong, H.-T., Shang, W.-J., Xu, X.-M., Zhu, H.-Q., Yang, J.-R., and Hu, X.-P. 2015. Threshold microsclerotial inoculum for cotton Verticillium wilt determined through wet-sieving and real-time quantitative PCR. Phytopathology 105:220-229.