Genotyping-by-Sequencing Training, Support, and Sequencing

Goals / Objectives
Objectives: (1) Train USDA personnel sample preparation for genotyping-by-sequencing (GBS) and (2) support sequencing of insect samples to identify single nucleotide polymorphisms by GBS that differentiate mosquito populations collected during the North American Mosquito Project. The sequencing will be done on disease vectors, Aedes vexans and Culex tarsalis, the two most likely disease vectors of JEV and RVF if they are introduced to the United States and the current disease vectors of EEE, WEE, SLE, and WNV. We will also include outgroup species such as Culex quinquefaciatus and Culex pipiens, the other two major Culex disease vectors. By knowing the genetic relatedness of the various mosquito subpopulations, we can determine the interepidemic maintance cycle and extent of spread from mosquito migration in the absence of an epidemic. Furthermore, by knowing the abiotic characteristics of the extant geographic areas, we can predict the future distributions of the diseases with considerations to global climate change. These sequences are necessary for phylogenetic and population genetic insect studies to ultimately understand the insect distributions, their impact on agriculture, and the future distributions accounting for global climate change.

Project Methods
Scope of work: The Kansas State University (KSU) lab will train USDA personnel in sample preparation and support the insect sequencing. This includes 96 well sample preparations (extraction, digestion, ligation, etc. ) and ultimately sequencing the barcoded individual insects with their automated processes. Samples will be prepared for sequencing in the KSU lab and the sequence data transferred to the USDA for processing and phylogenetic and population genetic analysis.