Non Technical Summary
Nitrogen is a critical nutrient for plant growth and yield. While nitrogen supply has been facilitated through the Haber-Bosch process and fertilizer application, over-application of fertilizer has drastic ecological consequences. Furthermore, nitrogen dose and time of application can have diverseinfluences on multiple traits of relevance to tomato growers including yield, fruit bioactive compound content, disease and seed quality. Therefore, there is an urgent need for plants with increased nitrogen use efficiency. Little is known regarding the molecular players associated with nitrogen metabolism in tomatoes. We have identified five tomato genes that determine plant N metabolic gene responsiveness and will utilize innovative approaches, guided by the needs of US tomato growers, breeders and stakeholders, to breed plants with increased nitrogen use efficiency.conditions.

Classification

Knowledge Area (KA)	Subject of Investigation (SOI)	Field of Science (FOS)	Percent
102	1460	1080	100%

Knowledge Area
102 - Soil, Plant, Water, Nutrient Relationships;

Subject Of Investigation
1460 - Tomato;

Field Of Science
1080 - Genetics;

Goals / Objectives
Aim 1: Test the predictive genetic model for five transcription factors (TF), their contribution to tomato NUE and their impact on tomato agronomic traitsAim 2: Investigate how these TFs regulate each other and refine the genetic model with targeted perturbations to increase transcriptional responsiveness to reduced levels of nitrateAim 3: Tune the dynamic range of these five TFs and their targets to engineer tomato plants with increased responsiveness to nitrogen (N) availability

Project Methods
Genotyping using amplicon-based sequencing.Phentoyping: plant leaf number, plant height, chlorophyll content, in situ level photosynthetic rate and stomatal conductance, fruit number, weight (fresh and dry), color, Brix, seed germination, fruit and seed size, shape, color and anatomy, protein content, 15N and 13C content.Statistical analyses: ANOVADetermination of regulatory dynamics: PAROT assay (protoplast-based assay with nitrogen reporter), in vitro identification of tomato TF-binding site data, TARGET assays for SlARF9B, SlARF18, SlNLP7A, SlNLP7B and SlDREB26 in limiting and sufficient N over timeGenetics: Manipulation of target genes in -cis and in -transGeneration of stable transgenic lines