Document Type

Thesis

Degree

Master of Science

Major

Biology

Date of Defense

9-4-2009

Graduate Advisor

Wendy M. Olivas, PhD

Co-Advisor

Claude M. Fauquet, Ph.D.

Committee

Sam X. Wang, Ph.D.

Bethany K. Zolman, Ph.D.

Abstract

Cassava (Manihot esculenta Crantz) is an important staple and cash crop in Africa, Latin America and Asia. In east and southern Africa, cassava brown streak disease (CBSD) caused by cassava brown streak virus (CBSV) is associated with significant losses in cassava production. Previously, the disease was prevalent only along coastal eastern and southern Africa, but it recently emerged in Uganda and is spreading rapidly in the country as well as in neighboring countries. Apart from a few cultivars that have shown tolerance to CBSV in Tanzania, no effective resistance to CBSV has been developed and deployed to date. The full genome sequence of CBSV is not yet known, but it is thought to be monopartite, linear, positive sense ssRNA, translated into a polyprotein that is further auto-cleaved into functional proteins with the capsid protein (CP) at the C-terminus. The present study aims to develop transient resistance to CBSV through CP-mediated protection and RNA interference (RNAi) strategies. The entire CBSV CP gene was used to express the CP and thereby trigger CP-mediated protection against CBSV. In addition, the full-length CP gene and its N- and C-terminal regions were used to generate three RNAi constructs, with RNAi-GFP as an internal control for transient studies in sap-inoculated GFP transgenic Nicotiana benthamiana. An efficient protocol for sap transmission of CBSV to N. benthamiana was also developed and used in transient protection studies of the constructs as proof-of-concept for control of CBSV using virus-derived resistance strategies in cassava. The constructs offered high levels of protection against CBSV and are highly recommended for use to transform cassava to generate CBSV resistant cassava plants for the farmers.

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