Date of Award

12-2019

Document Type

Thesis

Degree Name

Master of Science (MS)

Department

Biology

First Advisor

Seema Bharathan

Second Advisor

Narayanaswamy Bharathan

Third Advisor

Robert Major

Abstract

The soilborne, plant pathogenic fungus Rhizoctonia solani is classified into genetically isolated Anastomosis Groups (AGs). Within some of these AGs, Subgroups have been identified based on colony morphology host range, virulence, genetic sequences, and biochemical characteristics. Increased genetic variability is present between these groups. Present within R. solani are double-stranded (ds) RNA mycoviruses. These dsRNA viruses are genetically diverse both within and between fungal isolates. Plasmid DNA containing previously cloned dsRNA from AG 13 isolates RB 375, RB 303, & RB 386 was isolated, digested with EcoRI and analyzed via Gel Electrophoresis. The cloned dsRNA Fragments ranged from 0.764-1.88 kb. Real-time PCR was optimized with primers specific to the viral sequences. Viruses from all three isolates were detected using qPCR and annealing temperature of the primers was optimized at 52 °C with 98.32% efficiency. qPCR and phylogenetic analysis showed distinct genetic variations between viruses belonging to isolate 357 and between 357, 303, & 386. The bacterial housing these cloned viruses were edited using Clustered Regularly Interspaced Short Palindromic Repeats (CRISPR) selecting for Kanamycin resistance.

Share

COinS