Wendy A Harwood

Wendy A Harwood
John Innes Centre · Department of Crop Genetics

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124
Publications
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Publications

Publications (124)
Article
Full-text available
The granule-bound starch synthase I (GBSSI) encoded by the waxy gene is responsible for amylose synthesis in the endosperm of wheat grains. In the present study, a novel Wx-B1 null mutant line, M3-415, was identified from an ethyl methanesulfonate-mutagenized population of Chinese tetraploid wheat landrace Jianyangailanmai (LM47). The gene sequence...
Preprint
We report the first successful, high efficiency use of Lb Cas12a in barley and describe the development and application of two novel Cas12a variants. In total we compared five coding sequence (CDS) variants including the two novel ones and two guide architectures over 5 different target genes using twenty different guides. We found large difference...
Preprint
We report highly efficient genome editing in Brassica species. We compare the efficiency of targeted mutagenesis using four Streptococcus pyogenes Cas9 ( Sp Cas9) systems in Brassica oleracea and Brassica napus over 3 target genes and five guides to identify two which show a striking improvement to our first published system (Lawrenson et al., 2015...
Article
Full-text available
The wild relatives and progenitors of wheat have been widely used as sources of disease resistance (R) genes. Molecular identification and characterization of these R genes facilitates their manipulation and tracking in breeding programmes. Here, we develop a reference-quality genome assembly of the wild diploid wheat relative Aegilops sharonensis...
Article
Full-text available
Aegilops tauschii, the diploid wild progenitor of the D subgenome of bread wheat, is a reservoir of genetic diversity for improving bread wheat performance and environmental resilience. Here we sequenced 242 Ae. tauschii accessions and compared them to the wheat D subgenome to characterize genomic diversity. We found that a rare lineage of Ae. taus...
Article
Full-text available
In this study, a range of barley allelic mutants lost ADPG binding structure of starch synthase IIa (SSIIa) were created through targeted mutagenesis of SSIIa by RNA-guided Cas9. The transcriptomic and qRT-PCR results showed the increased mRNA expression of HvGBSSI and the decreased HvSSIIa and HvSBEI levels in ssIIa mutant grains, which were consi...
Preprint
Full-text available
The wild relatives and progenitors of wheat have been widely used as sources of disease resistance ( R ) genes. Molecular identification and characterization of these R genes facilitates their manipulation and tracking in breeding programmes. We developed a reference-quality genome assembly of the wild diploid wheat relative Aegilops sharonensis an...
Article
Full-text available
The APETALA2/Ethylene-Responsive factor (AP2/ERF) gene family is a large plant-specific transcription factor family, which plays important roles in regulating plant growth and development. A role in starch synthesis is among the multiple functions of this family of transcription factors. Barley (Hordeum vulgare L.) is one of the most important cere...
Article
Full-text available
The APETALA2/Ethylene-Responsive factor (AP2/ERF) gene family is a large plant-specific transcription factor family, which plays important roles in regulating plant growth and development. A role in starch synthesis is among the multiple functions of this family of transcription factors. Barley (Hordeum vulgare L.) is one of the most important cere...
Article
Full-text available
Advances in the use of RNA-guided Cas9-based genome editing in plants have been rapid over the last few years. A desirable application of genome editing is gene targeting (GT), as it allows a wide range of precise modifications; however, this remains inefficient especially in key crop species. Here, we describe successful, heritable gene targeting...
Article
Discoveries in model plants grown under optimal conditions can provide important directions for crop improvement. However, it is important to verify whether results can be translated to crop plants grown in the field. In this study, we sought to study the role of MYB28 in the regulation of aliphatic glucosinolate (A-GSL) biosynthesis and associated...
Article
Full-text available
Wheat, though a key crop plant with considerable influence on world food security, has nonetheless trailed behind other major cereals in the advancement of gene transformation technology for its improvement. New breeding technologies such as genome editing allow precise DNA manipulation, but their potential is limited by low regeneration efficienci...
Article
Full-text available
The development and application of high precision genome editing tools such as programmable nucleases are set to revolutionize crop breeding and are already having a major impact on fundamental science. Clustered regularly interspaced short palindromic repeats (CRISPR), and its CRISPR‐associated protein (Cas), is a programmable RNA‐guided nuclease...
Preprint
Full-text available
Advances in the use of RNA-guided Cas9-based genome editing in plants have been rapid over the last few years. A desirable application of genome editing is gene targeting (GT), as it allows a wide range of precise modifications, however this remains inefficient especially in key crop species. Here we describe successful, heritable gene targeting in...
Preprint
Full-text available
Aegilops tauschii, the diploid wild progenitor of the D-subgenome of bread wheat, constitutes a reservoir of genetic diversity for improving bread wheat performance and environmental resilience. To better define and understand this diversity, we sequenced 242 Ae. tauschii accessions and compared them to the wheat D-subgenome. We characterized a rar...
Article
Full-text available
In a ruling on 25 July 2018, the Court of Justice of the European Union concluded that organisms obtained by means of techniques/methods of mutagenesis constitute GMOs in the sense of Directive 2001/18, and that organisms obtained by means of techniques/methods of directed mutagenesis are not excluded from the scope of the Directive. Following the...
Article
Full-text available
In the last 20 years, stem rust caused by the fungus Puccinia graminis f. sp. tritici (Pgt), has reemerged as a major threat to wheat and barley production in Africa and Europe. In contrast to wheat with 82 designated stem rust (Sr) resistance genes, barley’s genetic variation for stem rust resistance is very narrow with only ten resistance genes g...
Preprint
Full-text available
We sought to quantify the role of MYB28 in the regulation of aliphatic glucosinolate biosynthesis and associated sulphur metabolism in field-grown B. oleracea with the use of CRISPR-Cas9-mediated gene editing technology. We describe the first characterised myb28 knockout mutant in B. oleracea, and the first DEFRA-regulated and approved CRISPR field...
Article
In wheat/alien species hybrids, pairing homoeologous (Ph1) suppresses the pairing and recombination of wheat and alien chromosomes. Ph1b has been found to be the most effective allele in inducing meiotic homoeologous pairing/recombination and is widely used in gene introgression from wild Triticeae species into wheat. However, due to the poor agron...
Article
Full-text available
In barley (Hordeum vulgare L.), Agrobacterium-mediated transformation efficiency is highly dependent on genotype with very few cultivars being amenable to transformation. Golden Promise is the cultivar most widely used for barley transformation and developing embryos are the most common donor tissue. We tested whether barley mutants with abnormally...
Article
Full-text available
Understanding the function of genes within staple crops will accelerate crop improvement by allowing targeted breeding approaches. Despite their importance, a lack of genomic information and resources has hindered the functional characterisation of genes in major crops. The recent release of high-quality reference sequences for these crops underpin...
Article
Three EMS mutant wheat lines with varied mobility of Wx-A1 and Wx-B1 were identified. In the mutant lines M2-97-8 and M2-104-2, Wx-B1 showed slightly faster mobility compared to those of wild line, and Wx-A1 in the mutant line M2-101-1 has a greatly faster mobility, which co-migrates with Wx-D1. The results of gene sequencing indicated that SNP mut...
Article
Full-text available
Improving nitrogen use efficiency (NUE) is very important for crops throughout the world. Rice mainly utilizes ammonium as an N source, but it also has four NRT2 genes involved in nitrate transport. The OsNRT2.3b transporter is important for maintaining cellular pH under mixed N supplies. Overexpression of this transporter driven by a ubiquitin pro...
Article
Full-text available
The waxy gene (Wx-1) encodes the granule-bound starch synthase (GBSSI or waxy protein, Wx), which is the key enzyme that catalyzes amylose synthesis in wheat. In this study, tetraploid wheat (Triticum turgidum durum L.) was mutagenized by ethyl methanesulfonate, and M2 populations of 3474 plants were generated. The Wx-A1 orWx-B1 null mutants were i...
Article
Full-text available
In a ruling on 25 July 2018, the Court of Justice of the European Union concluded that organisms obtained by means of techniques/methods of mutagenesis constitute GMOs in the sense of Directive 2001/18, and that organisms obtained by means of techniques/methods of directed mutagenesis are not excluded from the scope of the Directive. Following the...
Preprint
Full-text available
In barley (Hordeum vulgare L.), Agrobacterium-mediated transformation efficiency is highly dependent on genotype with very few cultivars being amenable to transformation. Golden Promise is the cultivar most widely used for barley transformation and developing embryos are the most common donor tissue. We tested whether barley mutants with abnormally...
Preprint
Full-text available
To adapt to the challenges of climate change and the growing world population, it is vital to increase global crop production. Understanding the function of genes within staple crops will accelerate crop improvement by allowing targeted breeding approaches. Despite the importance of wheat, which provides 20 % of the calories consumed by humankind,...
Preprint
To adapt to the challenges of climate change and the growing world population, it is vital to increase global crop production. Understanding the function of genes within staple crops will accelerate crop improvement by allowing targeted breeding approaches. Despite the importance of wheat, which provides 20 % of the calories consumed by humankind,...
Article
In our previous work, a novel high-molecular-weight glutenin subunit (HMW-GS) with an extremely large molecular weight from Aegilops sharonensis was identified that may contribute to excellent wheat (Triticum aestivum) processing quality and increased dough strength, and we further generated HMW-GS homozygous lines by crossing. In this study, we cr...
Article
In this study, we successfully knock-out the d-hordein component of barley storage protein using RNA-guided Cas9. Mutation frequencies of 25% and 14% at two different target sites were obtained. Homozygous mutant plants that were T-DNA free were identified in the T1 generation. Barley grains without d-hordein proteins from T2 seeds showed a signifi...
Article
Full-text available
Background: Despite wheat being a worldwide staple, it is still considered the most difficult to transform out of the main cereal crops. Therefore, for the wheat research community, a freely available and effective wheat transformation system is still greatly needed. Results: We have developed and optimised a reproducible Agrobacterium-mediated...
Article
Full-text available
Phosphatidylinositol transfer protein (PITP) and phosphatidylinositol 4-kinase (PI4K) are very upstream regulatory elements of the phospholipid signaling pathway in the signal transduction network. Unlike in animal systems, their role in stress signaling is poorly understood in plants. To study this area, PITP- and PI4K-overexpressing transgenic ba...
Article
Full-text available
Key message An EMS-induced single-base mutation at a splice site caused abnormal RNA splicing and resulted in the gene inactivation and the lack of Wx-A1 protein in a wheat EMS mutant line. Abstract An EMS-mutagenized population was generated using common wheat cv. SM126 consisting of 10,600 M2 plants. One Wx-A1 null mutant was identified through...
Article
Full-text available
We investigated whether Cas9‐mediated mutagenesis of starch‐branching enzymes (SBEs) in tetraploid potatoes could generate tuber starches with a range of distinct properties. Constructs containing the Cas9 gene and sgRNAs targeting SBE1, SBE2 or both genes were introduced by Agrobacterium‐mediated transformation or by PEG‐mediated delivery into pro...
Article
Full-text available
In this study, we successfully expressed the active 1Ay subunit of Triticum urartu in barley by Agrobacterium-mediated transformation with a transformation efficiency of 19.9%. The results of SDS–PAGE revealed that the expressed proteins of 1Ay subunit were present at some grains of each of 46 original T0 plants, showing identical mobility to those...
Article
Full-text available
Plant synthetic biology and cereal engineering depends on the controlled expression of transgenes of interest. Most engineering in plant species to date has relied heavily on the use of a few, well‐established constitutive promoters to achieve high levels of expression; however, the levels of transgene expression can also be influenced by the use o...
Chapter
Knockout mutants are an invaluable reverse genetics tool which have not been well developed in crop species compared to models like Arabidopsis. However, the emergence of CRISPR/Cas9 has changed this situation making the generation of such mutants accessible to many crops including barley. A single T-DNA construct can be transformed into barley imm...
Chapter
Barley has a number of unique features among our crop plants. It was one of the earliest plants to be domesticated and continues to play an important role in modern agriculture today. It is a versatile crop, used both for human nutrition and for animal feed, and plays an important role as an experimental model plant allowing advances in plant genet...
Chapter
Barley transformation is an essential tool for a range of functional genomics studies as well as for future crop improvement. The demand for efficient crop transformation systems continues to grow, with new genome editing technologies adding to that demand. Here we describe an efficient and routine transformation protocol for the spring barley Gold...
Chapter
While public and political views on genetic modification (inserting “foreign” genes to elicit new traits) have resulted in limited exploitation of the technology in some parts of the world, the new era of genome editing (to edit existing genes to gain new traits/genetic variation) has the potential to change the biotech landscape. Genome editing of...
Article
Full-text available
‘Speed breeding’ (SB) shortens the breeding cycle and accelerates crop research through rapid generation advancement. SB can be carried out in numerous ways, one of which involves extending the duration of plants’ daily exposure to light, combined with early seed harvest, to cycle quickly from seed to seed, thereby reducing the generation times for...
Article
Full-text available
Background Genetic transformation is a valuable tool and an important procedure in plant functional genomics contributing to gene discovery, allowing powerful insights into gene function and genetically controlled characteristics. Primulaceae species provide one of the best-known examples of heteromorphic flower development, a breeding system which...
Article
Full-text available
Dehydrins (Dhns) are the Group II Late Embryogenesis Abundant (LEA) proteins that are thought to play a key role in the response to abiotic stresses in plants. In the present study, a dehydrin6 (Dhn6) gene was isolated and characterized from eight barley genotypes (Hordeum vulgare L.) of diverse geographic origins, representing a wide range of drou...
Preprint
Full-text available
In the last 20 years, stem rust caused by the fungus Puccinia graminis f. sp. tritici ( Pgt ), has re-emerged as a major threat to wheat and barley cultivation in Africa and Europe. In contrast to wheat with 82 designated stem rust ( Sr ) resistance genes, barley's genetic variation for stem rust resistance is very narrow with only seven resistance...
Preprint
Full-text available
To meet the challenge of feeding a growing population, breeders and scientists are continuously looking for ways to increase genetic gain in crop breeding. One way this can be achieved is through 'speed breeding' (SB), which shortens the breeding cycle and accelerates research studies through rapid generation advancement. The SB method can be carri...
Conference Paper
Full-text available
The SNARE protein equivalent to the Arabidopsis SYP121 (also known as PEN1 and SYR1) is a member of the SNARE superfamily of proteins that are involved in cell signalling, vesicle traffic, growth and development. Key residues are F9-x-R11-F12, that form the core binding site for SM and Kv channel proteins. Eliminating this site dramatically alters...
Article
Full-text available
Is the European Union (EU) regulatory framework for genetically modified organisms (GMOs) adequate for emerging techniques, such as genome editing? This has been discussed extensively for more than 10 years. A recent proposal from The Netherlands offers a way to break the deadlock. Here, we discuss how the proposal would affect examples from public...
Data
Chromosomal configurations with single chiasma or double chiasmata highlighted with arrows. These structures marked by an arrow were counted as either single or double chiasmata in all analyzed meiocytes. Both datasets are shown in all analyzed genotypes. Bar: 20 μm.
Data
Representative meiotic configurations of Triticum aestivum cv. Cadenza (Cad1691-Tazip4-B2 mutant)-Ae. variabilis (A) and Triticum aestivum cv. Cadenza (Cad0348-Tazip4-B2 mutant)-Ae. variabilis (B) and wheat Tazip4-B2 CRISPR mutant-Ae. variabilis mutant (C) hybrids. From left to right: water alone, treated with either 1 mM Mg2+ or Hoagland solution....
Data
Genotypes and number of plants used for analyzing the effect of a nutrient solution in homoeologous CO frequency in wheat and its relative species.
Data
Frequencies of univalents, bivalents, multivalents and chiasma frequency (single and double chiasmata) were scored at meiotic metaphase I in wheat Tazip4-B2 CRISPR mutant-Ae. variabilis hybrids. Values in parenthesis indicate range of variation between cells. P < 0.05 indicates significant differences according to Dunn's test. Different letters ind...
Article
Full-text available
Wild relatives provide an important source of useful traits in wheat breeding. Wheat and wild relative hybrids have been widely used in breeding programs to introduce such traits into wheat. However, successful introgression is limited by the low frequency of homoeologous crossover (CO) between wheat and wild relative chromosomes. Hybrids between w...
Preprint
Full-text available
Wild relatives provide an important source of useful traits in wheat breeding. Wheat and wild relative hybrids have been widely used in breeding programs to introduce such traits into wheat. However, successful introgression is limited by the low frequency of homoeologous crossover (CO) between wheat and wild relative chromosomes. Hybrids between w...
Article
Full-text available
The growing human population and a changing environment have raised significant concern for global food security, with the current improvement rate of several important crops inadequate to meet future demand 1 . This slow improvement rate is attributed partly to the long generation times of crop plants. Here, we present a method called 'speed breed...
Article
Full-text available
Initially discovered in bacteria and archaea, CRISPR–Cas9 is an adaptive immune system found in prokaryotes. In 2012, scientists found a way to use it as a genome editing tool. In 2013, its application in plants was successfully achieved. This breakthrough has opened up many new opportunities for researchers, including the opportunity to gain a bet...