Location: Genetic Improvement for Fruits & Vegetables Laboratory
Project Number: 8042-21000-023-012-S
Project Type: Non-Assistance Cooperative Agreement
Start Date: Sep 1, 2022
End Date: Aug 31, 2025
Objective:
Research at ARS has developed solid genomic resources for cranberry (Vaccinium macrocarpon) including a reference quality genome assembly, genetic maps, genotyping and phenotyping protocols, etc. Currently lacking are methods to test and verify the function(s) of candidate genes associated with traits of interest. This issue will be addressed through two different objectives:
1. Development of an effective virus-induced gene silencing (VIGs) system for cranberry
2. Development of a CRISPR-Cas9 system for gene editing in cranberry
Approach:
1. A VIGs vector will be developed from blueberry scorch virus (BBScV) that is designed to deliver RNAi for gene silencing in cranberry. The vector will contain the RNA polymerase and coat protein coding genes from BBScV as well as a cloning site for inserting gene targets sequences.
2. ARS will construct our own CRISPR-editing vector that consists the following components: PAtU6::gRNA//PAtUbi::Cas9/TAtUbi. ARS will use the gRNA/Cas9 cassettes in the cloning vector pGEM3Z as a transient CRISPR-gene editing vector. We will use the gRNA/Cas9 cassettes in the plant transformation/expression vector pCAMBIA1300 as an integrating CRISPR-gene editing vector.
