Investigating the Role of CaML1-6 in Symbiotic Nitrogen Fixation via CRISPR-Cas9 Gene Knockouts in Medicago truncatula

Presenter

Skye Arielle McMorris

Campus

UMass Amherst

Sponsor

Dong Wang, Department of Biochemistry and Molecular Biology, UMass Amherst

Schedule

Session 1, 10:30 AM - 11:15 AM [Schedule by Time][Poster Grid for Time/Location]

Location

Poster Board A55, Campus Center Auditorium, Row 3 (A41-A60) [Poster Location Map]

Abstract

Legumes are afforded a hallmark of agricultural sustainability in that they can fix their own nitrogen via the symbiosis that occurs with soil bacteria known as rhizobia residing in their root cells. Calmodulin-like (CaML) calcium-binding proteins have been found to be specifically expressed in the root nodules where the symbionts live, but their function in the symbiosis remains unknown. In this study, genetic knockout of the 6 tandem CaML genes in the model legume M. truncatula using the genome-editing tool CRISPR-Cas9 will be generated in order to determine the role of these genes in the nitrogen-fixing symbiosis. Given the expression profile of the CaML genes, we hypothesize that this cluster is essential for the nitrogen fixation process in the symbiotic relationship, and thus knocking out these genes will prevent nitrogen fixation from occurring in the root nodules. Sequencing results of the targeted region used to genotype the mutants indicates that the construct is able to cause mutagenesis in the CaML genes and in low occurrence, delete the entire cluster. Future work aims to determine if there is a phenotype of M. truncatula that correlates with the deletion of the 6 CaML genes. Understanding the genetic basis of the nitrogen fixing symbiosis is a key to transferring this sustainable attribute to other non-nitrogen-fixing crops, thereby improving agricultural sustainability.

Keywords

legumes, rhizobia, nitrogen fixation, symbiosis, CRISPR-Cas9

Research Area

Agriculture and Agronomy / Food Science

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