Date of Graduation

7-2021

Document Type

Dissertation

Degree Name

Doctor of Philosophy in Plant Science (PhD)

Degree Level

Graduate

Department

Agricultural, Food and Life Sciences

Advisor/Mentor

Ainong Shi

Committee Member

Andy Pereira

Second Committee Member

Larry C. Purcell

Third Committee Member

Burt H. Bluhm

Fourth Committee Member

Vibha Srivastava

Keywords

bacterial wilt resistance, breeding, gene expression, genetics, pathogens, plant diseases, resistance, Spinach

Abstract

Spinach (Spinacia oleracea) is a self-pollinated, dioecious winter crop. Prevalent challenges to the production of spinach include disease pressure imposed by downy mildew, which is caused by Peronospora effusa (=P. farinosa f. sp. spinaciae [Pfs]). A total of 19 new races of P. effusa have emerged, imposing serious challenges to the disease management in spinach production. Accordingly, this study was designed to explore the genetic components for establishing the basis of durable disease resistance development against the downy mildew pathogen (P. effusa 13) in spinach, through the use of various genome engineering approaches.

Our results have led (Chapter 2) to the identification of potential stage specific resistance genes on the basis of the differential gene expression analysis between the downy mildew susceptible cultivar Viroflay and its near isogenic line NIL1 carrying downy mildew resistance gene RPF1. These genes included several novel genes encoding the NBS-LRR, RLK, PR- proteins along with the hormone associated genes, located on chromosomes 1, 3, 4, 5 and 6. Furthermore, the outcomes of this study propose a potential model for plant-pathogen interaction for the downy mildew infection in spinach by determining the stage specific resistance and susceptibility responses.

The IsoSeq studies identified the potential genes encoding the RLK, RLP, PR- protein and additional WRKY and NAC domain proteins localized on chromosomes 1, 3, 4, 5 and 6 specific to the downy mildew infection in Viroflay and NIL1 (Chapter 3). Further, relatively new genes were identified for the disease infection at the initial or control (0 hours post infection) and late infection or stress stages (48 hours post infection). Additionally, the full-length transcripts of the resistant and susceptible genotypes were analyzed by using two distinct spinach genome assemblies in order to elucidate the potential differences between the genomes of the two spinach genotypes, respectively.

Furthermore, a broader approach of gene pyramiding was used to stack the three Resistance to Peronospora farinosa (RPF) loci namely, RPF1, RPF2, and RPF3 resulting in stacking of three loci in a single spinach line and two loci in fifteen spinach lines. These findings will thus be useful in combatting multiple races of P. effusa, simultaneously (Chapter 4).

Overall, our study has facilitated the broadening of the genetic basis for plant pathogen interaction specific to the P. effusa infection, which causes downy mildew in spinach. It provides remarkable value to the scientific community by: 1) increased understanding of the stage specific dynamics of downy mildew infection by determining resistance or susceptibility loci in spinach genome, 2) clarifying the disparities of using two different genome assemblies in spinach, 3) identifying potentially new genetic elements, thereby broadening the knowledge of existing chromosomal location of disease resistance loci in spinach, and 4) establishing durable resistance by assembling three resistance loci into a single spinach line. Furthermore, the newly identified genetic elements associated with resistance to the downy mildew pathogen during this study can be used to further study and breed disease resistance cultivars of spinach.

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