2012 Annual Report 1a.Objectives (from AD-416): To understand the genetic complexity of field populations of the emerging species of potato cyst nematode (PCN) and to develop novel control tools to aid the eradication of PCN in Idaho. Specific objectives include: 1. Use the chorismate mutase gene as a genetic marker to evaluate genetic variations among G. pallida field populations. 2. Develop novel nematode resistance in potato through a plant-delivered RNAi technology. 1b.Approach (from AD-416): Globodera pallida, one of the two species of potato cyst nematode (PCN), has been found in nine fields in the eastern part of the state of Idaho. The discovery of this devastating potato pest poses threat to the potato industry of the U.S. Understanding the genetic complexity of field populations of this emerging nematode species and developing novel control tools are necessary to aid the eradication of G. pallida in Idaho. We will use the chorismate mutase gene as a genetic marker to evaluate genetic variations among G. pallida field populations; and, we will develop novel nematode resistance in potato through a plant-delivered RNAi technology. 3.Progress Report: The majority of research efforts focused on propagating PCN (G. pallida) field populations due to limited numbers of nematode cysts received from Idaho. ARS researchers have bulked up nematode cysts from three field populations and cloned the chorismate mutase (CM) gene from these nematode populations. Sequence analysis revealed that these three field populations contained one predominant GpCM sequence/allele, however, additional sequences/alleles were also identified. Results suggested that genetic variations exist among Idaho field populations. The plan is to clone the GpCM gene from other field populations to have a better understanding of the genetic diversity of G. pallida from Idaho. ARS researchers also cloned the CLE1 parasitism gene from G. pallida. Sequence analysis revealed that both CLE1 and CM genes are relatively conserved between the two PCN (G. rostochiensis and G. pallida) species. RNAi constructs were made targeting the conserved regions of these genes and transformed them into potato. Several transgenic potato lines expressing each of the RNAi construct were obtained. qRT-PCR assays confirmed the expression of dsRNA in the obtained transgenic potato lines. These transgenic potato lines will be tested for infection of both PCN species.