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Genome editing in Wheat

Ranjeet Kaur, Kshitija Sinha, Rupam Kumar Bhunia
Wheat is an important cereal crop that fulfils the calorie demands of the global humanity. Rapidly expanding populations are exposed to a fast approaching acute shortage in the adequate supply of food and fibre from agricultural resources. One of the significant threats to food security lies in the constantly increasing global temperatures which inflict serious injuries to the plants in terms of various physiological, biochemical and molecular processes. Wheat being a cool season crop is majorly impacted by the heat stress which adversely affects crop productivity and yield...
February 13, 2019: Molecular Biology Reports
Rakesh Kumar, Harohalli Masthigowda Mamrutha, Amandeep Kaur, Karnam Venkatesh, Davinder Sharma, Gyanendra Pratap Singh
Wheat is the most widely grown staple food crop in the world and accounts for dietary needs of more than 35% of the human population. Current status of transgenic wheat development is slow all over the world due to the lack of a suitable transformation system. In the present study, an efficient and reproducible Agrobacterium-mediated transformation system in bread wheat (Triticum aestivum L.) is established. The mature and immature embryos of six recently released high yielding spring bread wheat genotypes were used to standardize various parameters using Agrobacterium tumefaciens strain EHA105 harbouring binary vector pCAMBIA3301 having gus and bar as marker genes...
February 1, 2019: Molecular Biology Reports
Zhengzhi Zhang, Lei Hua, Ajay Gupta, David Tricoli, Keith J Edwards, Bing Yang, Wanlong Li
CRISPR/Cas9 has been widely used for genome editing in many organisms, including important crops like wheat. Despite the tractability in designing CRISPR/Cas9, efficacy in the application of this powerful genome editing tool also depends on DNA delivery methods. In wheat, the biolistics based transformation is the most used method for delivery of the CRISPR/Cas9 complex. Due to the high frequency of gene silencing associated with high copy transgenes and low edit rate in wheat, a large T0 transgenic plant population are required for recovery of desired mutations, which poses a bottleneck for many genome editing projects...
February 1, 2019: Plant Biotechnology Journal
Muhammad Amjad Ali, Mahpara Shahzadi, Adil Zahoor, Abdelfattah A Dababat, Halil Toktay, Allah Bakhsh, Muhammad Azher Nawaz, Hongjie Li
Cereal cyst nematodes (CCNs) are among the most important nematode pests that limit production of small grain cereals like wheat and barley. These nematodes alone are estimated to reduce production of crops by 10% globally. This necessitates a huge enhancement of nematode resistance in cereal crops against CCNs. Nematode resistance in wheat and barley in combination with higher grain yields has been a preferential research area for cereal nematologists. This usually involved the targeted genetic exploitations through natural means of classical selection breeding of resistant genotypes and finding quantitative trait luci (QTLs) associated with resistance genes...
January 19, 2019: International Journal of Molecular Sciences
Guoquan Liu, Jieqing Li, Ian D Godwin
The advanced biotechnology CRISPR/Cas9 as a powerful genome editing tool has displayed great potential for improving important agronomic traits such as yield and quality. It has gained momentum worldwide for gene function research of plants in recent years. As for cereals, numerous studies of CRISPR/Cas9 have been reported predominately on rice and quite a few on other cereals including maize, wheat, and barley. In contrast, there are only a couple of reports on sorghum up to date. In this chapter, the CRISPR/Cas9 system has been investigated for sorghum genome editing through biolistic bombardment...
2019: Methods in Molecular Biology
Wei Wang, Qianli Pan, Fei He, Alina Akhunova, Shiaoman Chao, Harold Trick, Eduard Akhunov
The CRISPR-Cas9-based multiplexed gene editing (MGE) provides a powerful method to modify multiple genomic regions simultaneously controlling different agronomic traits in crops. We applied the MGE construct built by combining the tandemly arrayed tRNA-gRNA units to generate heritable mutations in the TaGW2 , TaLpx-1 , and TaMLO genes of hexaploid wheat. The knockout mutations generated by this construct in all three homoeologous copies of one of the target genes, TaGW2 , resulted in a substantial increase in seed size and thousand grain weight...
February 1, 2018: The CRISPR journal
Shujuan Zhang, Rongzhi Zhang, Guoqi Song, Jie Gao, Wei Li, Xiaodong Han, Mingli Chen, Yulian Li, Genying Li
BACKGROUND: Recently, the CRISPR/Cas9 system has been widely used to precisely edit plant genomes. Due to the difficulty in Agrobacterium-mediated genetic transformation of wheat, the reported applications in CRISPR/Cas9 system were all based on the biolistic transformation. RESULTS: In the present study, we efficiently applied targeted mutagenesis in common wheat (Triticum aestivum L.) protoplasts and transgenic T0 plants using the CRISPR/Cas9 system delivered via Agrobacterium tumefaciens...
November 26, 2018: BMC Plant Biology
Qing Liu, Chun Wang, Xiaozhen Jiao, Huawei Zhang, Lili Song, Yanxin Li, Caixia Gao, Kejian Wang
The CRISPR/Cas system has been extensively applied to make precise genetic modifications in various organisms. Despite its importance and widespread use, large-scale mutation screening remains time-consuming, labour-intensive and costly. Here, we developed Hi-TOM (available at ), an online tool to track the mutations with precise percentage for multiple samples and multiple target sites. We also described a corresponding next-generation sequencing (NGS) library construction strategy by fixing the bridge sequences and barcoding primers...
November 13, 2018: Science China. Life Sciences
Philippa Borrill, Sophie A Harrington, Cristobal Uauy
Improving traits in wheat has historically been challenging due to its large and polyploid genome, limited genetic diversity, and in-field phenotyping constraints. However, within recent years many of these barriers have been lowered. The availability of a chromosome-level assembly of the wheat genome now facilitates a step-change in wheat genetics and provides a common platform for resources including variation data, gene expression data, and genetic markers. The development of sequenced mutant populations and gene-editing techniques now enable the rapid assessment of gene function in wheat directly...
November 8, 2018: Plant Journal: for Cell and Molecular Biology
Dmitry Miroshnichenko, Danila Ashin, Alexander Pushin, Sergey Dolgov
BACKGROUND: Domesticated einkorn (Triticum monococcum L.) is one of the oldest cultivated cereal crops in the world. Its small genome size (~ 5.7 GB), low ploidy (2n = 2x = 14, Am Am ) and high genetic polymorphism make this species very attractive for use as a diploid model for understanding the genomics and proteomics of Triticeae. Einkorn, however, is still a recalcitrant monocotyledonous species for the application of modern biotechnologies, including transgenesis. This paper reports the factors that may influence transgene delivery, integration, expression and inheritance in einkorn...
October 23, 2018: BMC Biotechnology
Zhuyun Bian, Yajia Ni, Jin-Rong Xu, Huiquan Liu
A-to-I RNA editing is an important post-transcriptional modification that converts adenosine (A) to inosine (I) in RNA molecules via hydrolytic deamination. Although editing of mRNAs catalyzed by adenosine deaminases acting on RNA (ADARs) is an evolutionarily conserved mechanism in metazoans, organisms outside the animal kingdom lacking ADAR orthologs were thought to lack A-to-I mRNA editing. However, recent discoveries of genome-wide A-to-I mRNA editing during the sexual stage of the wheat scab fungus Fusarium graminearum, model filamentous fungus Neurospora crassa, Sordaria macrospora, and an early diverging filamentous ascomycete Pyronema confluens indicated that A-to-I mRNA editing is likely an evolutionarily conserved feature in filamentous ascomycetes...
October 9, 2018: Cellular and Molecular Life Sciences: CMLS
Rhian M Howells, Melanie Craze, Sarah Bowden, Emma J Wallington
BACKGROUND: The use of CRISPR/Cas9 systems could prove to be a valuable tool in crop research, providing the ability to fully knockout gene function in complex genomes or to precisely adjust gene function by knockout of individual alleles. RESULTS: We compare gene editing in hexaploid wheat (Triticum aestivum) with diploid barley (Hordeum vulgare), using a combination of single genome and tri-genome targeting. High efficiency gene editing, 11-17% for single genome targeted guides and 5% for tri-genome targeted guides, was achieved in wheat using stable Agrobacterium-mediated transformation...
October 3, 2018: BMC Plant Biology
Haruyasu Hamada, Yuelin Liu, Yozo Nagira, Ryuji Miki, Naoaki Taoka, Ryozo Imai
The current application of genome editing to crop plants is limited to cultivars that are amenable to in vitro culture and regeneration. Here, we report an in planta genome-editing which does not require callus culture and regeneration. Shoot apical meristems (SAMs) contain a subepidermal cell layer, L2, from which germ cells later develop during floral organogenesis. The biolistic delivery of gold particles coated with plasmids expressing CRISPR/Cas9 components designed to target TaGASR7 were bombarded into SAM-exposed embryos of imbibed seeds...
September 26, 2018: Scientific Reports
Christian Zörb, Uwe Ludewig, Malcolm J Hawkesford
Wheat is an important cereal crop with a high demand for nitrogen (N) fertilizer to enable the grain protein accumulation that is necessary for baking and processing quality. Here, perspectives for the development of improved wheat genotypes with higher yield stability, better grain quality, and improved N use efficiency to lower environmental impacts are discussed. The development of improved wheat genotypes, for example, genotypes that lack storage proteins that do not contribute to baking quality (e.g., by genome editing), in combination with appropriate N fertilizer management to prevent N losses into the environment underpins a novel approach to improving N use efficiency...
November 2018: Trends in Plant Science
Virginia M G Borrelli, Vittoria Brambilla, Peter Rogowsky, Adriano Marocco, Alessandra Lanubile
Genome editing technologies have progressed rapidly and become one of the most important genetic tools in the implementation of pathogen resistance in plants. Recent years have witnessed the emergence of site directed modification methods using meganucleases, zinc finger nucleases (ZFNs), transcription activator-like effector nucleases (TALENs), and clustered regularly interspaced short palindrome repeats (CRISPR)/CRISPR-associated protein 9 (Cas9). Recently, CRISPR/Cas9 has largely overtaken the other genome editing technologies due to the fact that it is easier to design and implement, has a higher success rate, and is more versatile and less expensive...
2018: Frontiers in Plant Science
Wei Wang, James Simmonds, Qianli Pan, Dwight Davidson, Fei He, Abdulhamit Battal, Alina Akhunova, Harold N Trick, Cristobal Uauy, Eduard Akhunov
CRISPR-Cas9-based genome editing and EMS mutagenesis revealed inter-cultivar differences and additivity in the contribution of TaGW2 homoeologues to grain size and weight in wheat. The TaGW2 gene homoeologues have been reported to be negative regulators of grain size (GS) and thousand grain weight (TGW) in wheat. However, the contribution of each homoeologue to trait variation among different wheat cultivars is not well documented. We used the CRISPR-Cas9 system and TILLING to mutagenize each homoeologous gene copy in cultivars Bobwhite and Paragon, respectively...
November 2018: TAG. Theoretical and Applied Genetics. Theoretische und Angewandte Genetik
Aurélie Jouanin, Luud J W J Gilissen, Lesley A Boyd, James Cockram, Fiona J Leigh, Emma J Wallington, Hetty C van den Broeck, Ingrid M van der Meer, Jan G Schaart, Richard G F Visser, Marinus J M Smulders
A strict gluten-free diet is currently the only treatment for the 1-2% of the world population who suffer from coeliac disease (CD). However, due to the presence of wheat and wheat derivatives in many food products, avoiding gluten consumption is difficult. Gluten-free products, made without wheat, barley or rye, typically require the inclusion of numerous additives, resulting in products that are often less healthy than gluten-based equivalents. Here, we present and discuss two broad approaches to decrease wheat gluten immunogenicity for CD patients...
August 2018: Food Research International
Ajith Anand, Todd J Jones
The last decade has seen significant strides in Agrobacterium-mediated plant transformation technology. This has not only expanded the number of crop species that can be transformed by Agrobacterium, but has also made it possible to routinely transform several recalcitrant crop species including cereals (e.g., maize, sorghum, and wheat). However, the technology is limited by the random nature of DNA insertions, genotype dependency, low frequency of quality events, and variation in gene expression arising from genomic insertion sites...
June 30, 2018: Current Topics in Microbiology and Immunology
Yidong Ran, Nicola Patron, Pippa Kay, Debbie Wong, Margaret Buchanan, Ying-Ying Cao, Tim Sawbridge, John P Davies, John Mason, Steven R Webb, German Spangenberg, William M Ainley, Terence A Walsh, Matthew J Hayden
Sequence-specific nucleases have been used to engineer targeted genome modifications in various plants. While targeted gene knockouts resulting in loss of function have been reported with relatively high rates of success, targeted gene editing using an exogenously supplied DNA repair template and site-specific transgene integration has been more challenging. Here, we report the first application of zinc finger nuclease (ZFN)-mediated, nonhomologous end-joining (NHEJ)-directed editing of a native gene in allohexaploid bread wheat to introduce, via a supplied DNA repair template, a specific single amino acid change into the coding sequence of acetohydroxyacid synthase (AHAS) to confer resistance to imidazolinone herbicides...
December 2018: Plant Biotechnology Journal
Zhen Liang, Kunling Chen, Yan Yan, Yi Zhang, Caixia Gao
Despite the great achievements in genome editing, accurately detecting mutations induced by sequence-specific nucleases is still a challenge in plants, especially in polyploidy plants. An efficient detection method is particularly vital when the mutation frequency is low or when a large population needs to be screened. Here, we applied purified CRISPR ribonucleoprotein complexes to cleave PCR products for genome-edited mutation detection in hexaploid wheat and diploid rice. We show that this mutation detection method is more sensitive than Sanger sequencing and more applicable than PCR/RE method without the requirement for restriction enzyme site...
December 2018: Plant Biotechnology Journal
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