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Virus-induced genome editing in wheat
Thesis (MScAgric)--Stellenbosch University, 2025.
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Stellenbosch : Stellenbosch University
2025
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| _version_ | 1867613742592688128 |
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| access_status_str | Open Access |
| author | Bosman, Rudolph |
| author2 | Campa, Manuela |
| author_browse | Bosman, Rudolph Campa, Manuela |
| author_facet | Campa, Manuela Bosman, Rudolph |
| author_sort | Bosman, Rudolph |
| collection | Thesis |
| dc_rights_str_mv | Stellenbosch University |
| description | Thesis (MScAgric)--Stellenbosch University, 2025. |
| format | Thesis |
| id | oai:scholar.sun.ac.za:10019.1/132079 |
| institution | Stellenbosch University (South Africa) |
| last_indexed | 2026-06-10T12:40:58.715Z |
| license_str | Other — see source repository |
| provenance_str_mv | Harvested via OAI-PMH from SUNScholar — Stellenbosch University Repository |
| publishDate | 2025 |
| publishDateRange | 2025 |
| publishDateSort | 2025 |
| publisher | Stellenbosch : Stellenbosch University |
| publisherStr | Stellenbosch : Stellenbosch University |
| record_format | dspace |
| source_str | SUNScholar — Stellenbosch University Repository |
| spelling | oai:scholar.sun.ac.za:10019.1/132079 Virus-induced genome editing in wheat Bosman, Rudolph Campa, Manuela Burger, Johan Stellenbosch University. Faculty of AgriSciences. Dept. of Genetics. Wheat -- Genetic engineering Wheat -- Breeding Wheat -- Genome mapping Agrobacterium tumefaciens CRISPR (Genetics) Barley stripe mosaic virus Wheat -- Disease and pest resistance -- Genetic aspects UCTD Thesis (MScAgric)--Stellenbosch University, 2025. Bosman, R. 2025. Virus-induced genome editing in wheat. Unpublished masters thesis. Stellenbosch: Stellenbosch University [online]. Available: https://scholar.sun.ac.za/items/0a3a75b1-5c9a-4f85-9288-43af45ec2475 ENGLISH ABSTRACT: Bread wheat (Triticum aestivum L.) is the most widely grown staple crop in the world and is vitally important to maintain global food security. However, current rates of wheat production are inadequate to meet the food demands of the growing world population. The advent of genome editing technology such as clustered regularly-interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9) has revolutionised crop improvement due to the ability to efficiently and precisely induce mutations. However, wheat has lagged behind other crops in terms of genome editing due to its hexaploid genome and recalcitrance to tissue culture and regeneration. There have been recent advancements to address these challenges. To improve the low transformation efficiency of wheat, optimised Agrobacterium-mediated transformation protocols have been developed in recent years. Further, the overexpression of developmental regulatory genes was shown to increase the regeneration efficiency of wheat during tissue culture. Additionally, barley stripe mosaic virus (BSMV)-mediated single guide RNA (sgRNA) delivery has recently been used to efficiently induce genome editing in wheat. This study investigated the integration of these technologies with the goal of producing genome-edited wheat plants. Firstly, Agrobacterium-mediated transformation of immature embryos was combined with overexpression of wheat GROWTH REGULATING FACTOR 4 (GRF4) fused to its co-factor GRF-INTERACTING FACTOR 1 (GIF1). This resulted in the successful regeneration of Cas9-expressing wheat plants of the variety Fielder. Further, the transgene was shown to be inherited and expressed in subsequent generations. Thereafter, barley stripe mosaic virus was modified to deliver sgRNAs into Cas9-expressing wheat plants, with the aim of inducing genome editing in the gene phytoene desaturase. However, no edits could be detected, likely due to unsuccessful mechanical inoculations of the wheat plants with BSMV. Although genome editing was not achieved, this study provides general insights into the processes in both wheat transformations and virus-induced genome editing. Finally, this study explores potential optimisations and future prospects of utilising barley stripe mosaic virus to induce genome editing in wheat. AFRIKAANSE OPSOMMING: Broodkoring (Triticum aestivum L.) is die stapelgewas wat die meeste verbou word ter wêreld, en is van kritiese belang vir die instandhouding van globale voedselsekuriteit. Nieteenstaande is die huidige tempo van koringproduksie onvoldoende om aan die voedselbehoeftes van die groeiende wêreldbevolking te voldoen. Ontwikkelinge in genoomredigeringstegnologie soos ‘clustered regularly-interspaced short palindromic repeats (CRISPR)/CRISPR associated protein 9 (Cas9)’ het ’n groot omwenteling in gewasverbetering teweeggebring deur die tegnologie se vermoë om mutasies doeltreffend en akkuraat te induseer. Koring het egter wat genoomredigering betref agtergebly teenoor ander gewasse, weens die gewas se heksaploïede genoom en weerstandigheid teen weefselkultuur en regenerasie. Daar is egter onlangs vordering gemaak om hierdie uitdagings te bowe te kom. Oor die afgelope paar jaar is geoptimaliseerde Agrobacterium-bemiddelde transformasieprotokolle ontwikkel om die lae transformasiedoeltreffendheid van koring te verbeter. Verder is daar gedemonstreer dat die ooruitdrukking van ontwikkelingsregulerende gene die regenerasiedoeltreffendheid van koring tydens weefselkultuur verhoog. Barley stripe mosaic virus (BSMV)-bemiddelde aflewering van enkele gids-RNAs (sgRNAs) is onlangs suksesvol gebruik om genoomredigering in koring te bewerkstellig. Hierdie studie het die integrasie van die bostaande tegnologieë ondersoek met die doel om genoomgeredigeerde koringplante te produseer. Eerstens is Agrobacterium-bemiddelde transformasie van onvolwasse embrio’s gekombineer met die ooruitdrukking van koring GROWTH REGULATING FACTOR 4 (GRF4) wat aan sy ko-faktor GRF-INTERACTING FACTOR 1 (GIF1) gekoppel is. Dit het gelei tot die suksesvolle regenerasie van Cas9- uitdrukkende koringplante van die variëteit Fielder. Voorts is getoon dat die transgeen suksesvol na opeenvolgende generasies oorgedra en uitgedruk word. Daarna is BSMV gemodifiseer om sgRNA’s in Cas9-uitdrukkende koringplante af te lewer, met die doel om genoomredigering in die geen phytoene desaturase te induseer. Geenredigering kon egter nie bewys word nie, waarskynlik as gevolg van onsuksesvolle meganiese inokulasies van die koringplante met BSMV. Alhoewel genoomredigering nie plaasgevind het nie, bied hierdie studie waardevolle insigte in beide koringtransformasieprosesse en virus-geïnduseerde genoomredigering. Laastens word daar enkele voorstelle gemaak wat betref potensiële optimaliserings van en vooruitsigte vir die gebruik van barley stripe mosaic virus om genoomredigering in koring te bewerkstellig. Masters 2025-05-22T12:13:29Z 2025-05-22T12:13:29Z 2025-03 Thesis https://scholar.sun.ac.za/handle/10019.1/132079 Stellenbosch University xii, 79 pages : illustrations application/pdf Stellenbosch : Stellenbosch University |
| spellingShingle | Wheat -- Genetic engineering Wheat -- Breeding Wheat -- Genome mapping Agrobacterium tumefaciens CRISPR (Genetics) Barley stripe mosaic virus Wheat -- Disease and pest resistance -- Genetic aspects UCTD Bosman, Rudolph Virus-induced genome editing in wheat |
| title | Virus-induced genome editing in wheat |
| title_full | Virus-induced genome editing in wheat |
| title_fullStr | Virus-induced genome editing in wheat |
| title_full_unstemmed | Virus-induced genome editing in wheat |
| title_short | Virus-induced genome editing in wheat |
| title_sort | virus induced genome editing in wheat |
| topic | Wheat -- Genetic engineering Wheat -- Breeding Wheat -- Genome mapping Agrobacterium tumefaciens CRISPR (Genetics) Barley stripe mosaic virus Wheat -- Disease and pest resistance -- Genetic aspects UCTD |
| url | https://scholar.sun.ac.za/handle/10019.1/132079 |
| work_keys_str_mv | AT bosmanrudolph virusinducedgenomeeditinginwheat |