Simultaneous CRISPR/Cas9-mediated editing of cassava eIF4E isoforms nCBP-1 and nCBP-2 reduces cassava brown streak disease symptom severity and incidence

Plant Biotechnology Journal
Michael A GomezRebecca S Bart

Abstract

Cassava brown streak disease (CBSD) is a major constraint on cassava yields in East and Central Africa and threatens production in West Africa. CBSD is caused by two species of positive-sense RNA viruses belonging to the family Potyviridae, genus Ipomovirus: Cassava brown streak virus (CBSV) and Ugandan cassava brown streak virus (UCBSV). Diseases caused by the family Potyviridae require the interaction of viral genome-linked protein (VPg) and host eukaryotic translation initiation factor 4E (eIF4E) isoforms. Cassava encodes five eIF4E proteins: eIF4E, eIF(iso)4E-1, eIF(iso)4E-2, novel cap-binding protein-1 (nCBP-1), and nCBP-2. Protein-protein interaction experiments consistently found that VPg proteins associate with cassava nCBPs. CRISPR/Cas9-mediated genome editing was employed to generate ncbp-1, ncbp-2, and ncbp-1/ncbp-2 mutants in cassava cultivar 60444. Challenge with CBSV showed that ncbp-1/ncbp-2 mutants displayed delayed and attenuated CBSD aerial symptoms, as well as reduced severity and incidence of storage root necrosis. Suppressed disease symptoms were correlated with reduced virus titre in storage roots relative to wild-type controls. Our results demonstrate the ability to modify multiple genes simultaneously in...Continue Reading

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Methods Mentioned

BETA
transgenic
Immunoprecipitation
PCR

Software Mentioned

SnapGene
BioNJ
BLAST
Phytozome
MEGA
CRISPR
CasOT
Clustal Omega
CLC Genomics Workbench

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