Frontiers in genome editing最新文献

{"title":"Establishment of a genome editing tool using CRISPR-Cas9 ribonucleoprotein complexes in the non-model plant pathogen <i>Sphaerulina musiva</i>.","authors":"Joanna Tannous,&nbsp;Cole Sawyer,&nbsp;Md Mahmudul Hassan,&nbsp;Jesse L Labbe,&nbsp;Carrie Eckert","doi":"10.3389/fgeed.2023.1110279","DOIUrl":"https://doi.org/10.3389/fgeed.2023.1110279","url":null,"abstract":"<p><p>CRISPR-Cas9 is a versatile genome editing system widely used since 2013 to introduce site-specific modifications into the genomes of model and non-model species. This technology is used in various applications, from gene knock-outs, knock-ins, and over-expressions to more precise changes, such as the introduction of nucleotides at a targeted locus. CRISPR-Cas9 has been demonstrated to be easy to establish in new species and highly efficient and specific compared to previous gene editing strategies such as Zinc finger nucleases and transcription activator-like effector nucleases. Grand challenges for emerging CRISPR-Cas9 tools in filamentous fungi are developing efficient transformation methods for non-model organisms. In this paper, we have leveraged the establishment of CRISPR-Cas9 genome editing tool that relies on Cas9/sgRNA ribonucleoprotein complexes (RNPs) in the model species <i>Trichoderma reesei</i> and developed the first protocol to efficiently transform the non-model species, <i>Sphaerulina musiva.</i> This fungal pathogen constitutes a real threat to the genus <i>Populus,</i> a foundational bioenergy crop used for biofuel production. Herein, we highlight the general considerations to design sgRNAs and their computational validation. We also describe the use of isolated protoplasts to deliver the CRISPR-Cas9 RNP components in both species and the screening for targeted genome editing events. The development of engineering tools in <i>S. musiva</i> can be used for studying genes involved in diverse processes such as secondary metabolism, establishment, and pathogenicity, among many others, but also for developing genetic mitigation approaches. The approach described here provides guidance for potential development of transformation systems in other non-model spore-bearing ascomycetes.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2023-01-01","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC10401582/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10308295","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Efficient <i>hyperactive piggyBac</i> transgenesis in <i>Plodia</i> pantry moths.","authors":"Christa Heryanto, Anyi Mazo-Vargas, Arnaud Martin","doi":"10.3389/fgeed.2022.1074888","DOIUrl":"10.3389/fgeed.2022.1074888","url":null,"abstract":"<p><p>While <i>piggyBac</i> transposon-based transgenesis is widely used in various emerging model organisms, its relatively low transposition rate in butterflies and moths has hindered its use for routine genetic transformation in Lepidoptera. Here, we tested the suitability of a codon-optimized <i>hyperactive piggyBac</i> transposase (<i>hyPBase</i>) in mRNA form to deliver and integrate transgenic cassettes into the genome of the pantry moth <i>Plodia interpunctella</i>. Co-injection of <i>hyPBase</i> mRNA with donor plasmids successfully integrated 1.5-4.4 kb expression cassettes driving the fluorescent markers EGFP, DsRed, or EYFP in eyes and glia with the <i>3xP3</i> promoter. Somatic integration and expression of the transgene in the G₀ injected generation was detectable from 72-h embryos and onward in larvae, pupae and adults carrying a recessive white-eyed mutation. Overall, 2.5% of injected eggs survived into transgene-bearing adults with mosaic fluorescence. Subsequent outcrossing of fluorescent G₀ founders transmitted single-insertion copies of <i>3xP3::EGFP</i> and <i>3xP3::EYFP</i> and generated stable isogenic lines. Random in-crossing of a small cohort of G₀ founders expressing <i>3xP3::DsRed</i> yielded a stable transgenic line segregating for more than one transgene insertion site. We discuss how <i>hyPBase</i> can be used to generate stable transgenic resources in <i>Plodia</i> and other moths.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":4.9,"publicationDate":"2022-12-23","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9816379/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10874972","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Imaging tools for plant nanobiotechnology.","authors":"Bin Zhao, Zhongxu Luo, Honglu Zhang, Huan Zhang","doi":"10.3389/fgeed.2022.1029944","DOIUrl":"10.3389/fgeed.2022.1029944","url":null,"abstract":"<p><p>The successful application of nanobiotechnology in biomedicine has greatly changed the traditional way of diagnosis and treating of disease, and is promising for revolutionizing the traditional plant nanobiotechnology. Over the past few years, nanobiotechnology has increasingly expanded into plant research area. Nanomaterials can be designed as vectors for targeted delivery and controlled release of fertilizers, pesticides, herbicides, nucleotides, proteins, etc. Interestingly, nanomaterials with unique physical and chemical properties can directly affect plant growth and development; improve plant resistance to disease and stress; design as sensors in plant biology; and even be used for plant genetic engineering. Similarly, there have been concerns about the potential biological toxicity of nanomaterials. Selecting appropriate characterization methods will help understand how nanomaterials interact with plants and promote advances in plant nanobiotechnology. However, there are relatively few reviews of tools for characterizing nanomaterials in plant nanobiotechnology. In this review, we present relevant imaging tools that have been used in plant nanobiotechnology to monitor nanomaterial migration, interaction with and internalization into plants at three-dimensional lengths. Including: 1) Migration of nanomaterial into plant organs 2) Penetration of nanomaterial into plant tissues (iii)Internalization of nanomaterials by plant cells and interactions with plant subcellular structures. We compare the advantages and disadvantages of current characterization tools and propose future optimal characterization methods for plant nanobiotechnology.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":4.9,"publicationDate":"2022-12-08","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9772283/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10436908","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"CRISPR nuclease off-target activity and mitigation strategies.","authors":"Beeke Wienert, M Kyle Cromer","doi":"10.3389/fgeed.2022.1050507","DOIUrl":"10.3389/fgeed.2022.1050507","url":null,"abstract":"<p><p>The discovery of CRISPR has allowed site-specific genomic modification to become a reality and this technology is now being applied in a number of human clinical trials. While this technology has demonstrated impressive efficacy in the clinic to date, there remains the potential for unintended on- and off-target effects of CRISPR nuclease activity. A variety of <i>in silico</i>-based prediction tools and empirically derived experimental methods have been developed to identify the most common unintended effect-small insertions and deletions at genomic sites with homology to the guide RNA. However, large-scale aberrations have recently been reported such as translocations, inversions, deletions, and even chromothripsis. These are more difficult to detect using current workflows indicating a major unmet need in the field. In this review we summarize potential sequencing-based solutions that may be able to detect these large-scale effects even at low frequencies of occurrence. In addition, many of the current clinical trials using CRISPR involve <i>ex vivo</i> isolation of a patient's own stem cells, modification, and re-transplantation. However, there is growing interest in direct, <i>in vivo</i> delivery of genome editing tools. While this strategy has the potential to address disease in cell types that are not amenable to <i>ex vivo</i> manipulation, <i>in vivo</i> editing has only one desired outcome-on-target editing in the cell type of interest. CRISPR activity in unintended cell types (both on- and off-target) is therefore a major safety as well as ethical concern in tissues that could enable germline transmission. In this review, we have summarized the strengths and weaknesses of current editing and delivery tools and potential improvements to off-target and off-tissue CRISPR activity detection. We have also outlined potential mitigation strategies that will ensure that the safety of CRISPR keeps pace with efficacy, a necessary requirement if this technology is to realize its full translational potential.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":4.9,"publicationDate":"2022-11-10","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9685173/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"10658011","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Nanoparticles targeting hematopoietic stem and progenitor cells: Multimodal carriers for the treatment of hematological diseases.","authors":"Luis J Cruz,&nbsp;Somayeh Rezaei,&nbsp;Frank Grosveld,&nbsp;Sjaak Philipsen,&nbsp;Christina Eich","doi":"10.3389/fgeed.2022.1030285","DOIUrl":"https://doi.org/10.3389/fgeed.2022.1030285","url":null,"abstract":"<p><p>Modern-day hematopoietic stem cell (HSC) therapies, such as gene therapy, modify autologous HSCs prior to re-infusion into myelo-conditioned patients and hold great promise for treatment of hematological disorders. While this approach has been successful in numerous clinical trials, it relies on transplantation of <i>ex vivo</i> modified patient HSCs, which presents several limitations. It is a costly and time-consuming procedure, which includes only few patients so far, and <i>ex vivo</i> culturing negatively impacts on the viability and stem cell-properties of HSCs. If viral vectors are used, this carries the additional risk of insertional mutagenesis. A therapy delivered to HSCs <i>in vivo</i>, with minimal disturbance of the HSC niche, could offer great opportunities for novel treatments that aim to reverse disease symptoms for hematopoietic disorders and could bring safe, effective and affordable genetic therapies to all parts of the world. However, substantial unmet needs exist with respect to the <i>in vivo</i> delivery of therapeutics to HSCs. In the last decade, in particular with the development of gene editing technologies such as CRISPR/Cas9, nanoparticles (NPs) have become an emerging platform to facilitate the manipulation of cells and organs. By employing surface modification strategies, different types of NPs can be designed to target specific tissues and cell types <i>in vivo</i>. HSCs are particularly difficult to target due to the lack of unique cell surface markers that can be utilized for cell-specific delivery of therapeutics, and their shielded localization in the bone marrow (BM). Recent advances in NP technology and genetic engineering have resulted in the development of advanced nanocarriers that can deliver therapeutics and imaging agents to hematopoietic stem- and progenitor cells (HSPCs) in the BM niche. In this review we provide a comprehensive overview of NP-based approaches targeting HSPCs to control and monitor HSPC activity <i>in vitro</i> and <i>in vivo</i>, and we discuss the potential of NPs for the treatment of malignant and non-malignant hematological disorders, with a specific focus on the delivery of gene editing tools.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2022-11-02","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9666682/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40699729","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Plant biomacromolecule delivery methods in the 21st century.","authors":"Sachin Rustgi, Salman Naveed, Jonathan Windham, Huan Zhang, Gözde S Demirer","doi":"10.3389/fgeed.2022.1011934","DOIUrl":"10.3389/fgeed.2022.1011934","url":null,"abstract":"<p><p>The 21st century witnessed a boom in plant genomics and gene characterization studies through RNA interference and site-directed mutagenesis. Specifically, the last 15 years marked a rapid increase in discovering and implementing different genome editing techniques. Methods to deliver gene editing reagents have also attempted to keep pace with the discovery and implementation of gene editing tools in plants. As a result, various transient/stable, quick/lengthy, expensive (requiring specialized equipment)/inexpensive, and versatile/specific (species, developmental stage, or tissue) methods were developed. A brief account of these methods with emphasis on recent developments is provided in this review article. Additionally, the strengths and limitations of each method are listed to allow the reader to select the most appropriate method for their specific studies. Finally, a perspective for future developments and needs in this research area is presented.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":4.9,"publicationDate":"2022-10-14","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9614364/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"9176663","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"Environmental and socio-economic impacts of new plant breeding technologies: A case study of root chicory for inulin production.","authors":"Maria Hingsamer,&nbsp;Veronika Kulmer,&nbsp;Matthew de Roode,&nbsp;Michael Kernitzkyi","doi":"10.3389/fgeed.2022.919392","DOIUrl":"https://doi.org/10.3389/fgeed.2022.919392","url":null,"abstract":"<p><p>In Europe, root chicory and other plants are cultivated for their prebiotic food fiber, inulin, which boosts the growth of beneficial gut bacteria and stimulates the human immune system. CHIC, a H2020 project, develops new chicory variants which produce more and reported to be healthier inulin as well as medicinal terpenes. This paper presents an environmental and socio-economic assessment of the whole value chain of the new chicory variants and their derived products using a case study based in the Netherlands. Two scenarios based on new chicory variants using new plant breeding technologies (NPBT) are analyzed and impacts thereof are compared to the reference scenario; the current commercial inulin process from conventional chicory. Both scenarios show higher inulin content, but the inulin adsorption process differs. While one aims to optimize inulin yield, the other one explores the potential of a multipurpose use, yielding inulin and health beneficial terpenes. Methodologically, we employ multi-regional input-output (MRIO) analysis to estimate additional economic benefits, added value and job creation, while by means of life cycle assessment (LCA) effects on greenhouse gas (GHG) emissions and primary energy demand are derived. Both methods, MRIO and LCA, are well suited to analyze the raised issues and draw on the same data. Generally, the results highlight the importance of inulin production at a national and EU-level in the reference scenario. In case of the two scenarios, we find that the related socio-economic impacts are much higher than in the reference scenario and thus highlight their ability to boost economic activity and increase competiveness of the EU, i.e. over 80% of the generated value added stays in the EU. In terms of environmental impacts, the two scenarios show lower GHG emissions and primary energy demand due to the higher efficiencies of the process in the scenarios compared to the reference inulin process. Additionally, regarding the goal of climate neutral production, we find that the majority of GHG emissions stem from the electricity mix and natural gas demand. Replacing these sources of energy with more renewable ones will contribute to this goal.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2022-10-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9582860/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40652011","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"<i>In vivo</i> and <i>in vitro</i> genome editing to explore GNE functions.","authors":"Nili Ilouz,&nbsp;Avi Harazi,&nbsp;Miriam Guttman,&nbsp;Alon Daya,&nbsp;Shmuel Ruppo,&nbsp;Lena Yakovlev,&nbsp;Stella Mitrani-Rosenbaum","doi":"10.3389/fgeed.2022.930110","DOIUrl":"https://doi.org/10.3389/fgeed.2022.930110","url":null,"abstract":"<p><p>GNE myopathy is an adult onset neuromuscular disorder characterized by slowly progressive distal and proximal muscle weakness, caused by missense recessive mutations in the <i>GNE</i> gene. Although the encoded bifunctional enzyme is well known as the limiting factor in the biosynthesis of sialic acid, no clear mechanisms have been recognized to account for the muscle atrophic pathology, and novel functions for GNE have been hypothesized. Two major issues impair studies on this protein. First, the expression of the GNE protein is minimal in human and mice muscles and there is no reliable antibody to follow up endogenous expression. Second, no reliable animal model is available for the disease and cellular models from GNE myopathy patients' muscle cells (expressing the mutated protein) are less informative than expected. In order to broaden our knowledge on GNE functions in muscle, we have taken advantage of the CRISPR/Cas9 method for genome editing to first, add a tag to the endogenous Gne gene in mouse, allowing the determination of the spatiotemporal expression of the protein in the organism, using well established and reliable antibodies against the specific tag. In addition we have generated a Gne knock out murine muscle cell lineage to identify the events resulting from the total lack of the protein. A thorough multi-omics analysis of both cellular systems including transcriptomics, proteomics, phosphoproteomics and ubiquitination, unraveled novel pathways for Gne, in particular its involvement in cell cycle control and in the DNA damage/repair pathways. The elucidation of fundamental mechanisms of Gne in normal muscle may contribute to the identification of the disrupted functions in GNE myopathy, thus, to the definition of novel biomarkers and possible therapeutic targets for this disease.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2022-09-27","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9552322/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33509556","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"CRISPR/Cas genome editing improves abiotic and biotic stress tolerance of crops.","authors":"Yangyang Li,&nbsp;Xiuzhe Wu,&nbsp;Yan Zhang,&nbsp;Qiang Zhang","doi":"10.3389/fgeed.2022.987817","DOIUrl":"https://doi.org/10.3389/fgeed.2022.987817","url":null,"abstract":"<p><p>Abiotic stress such as cold, drought, saline-alkali stress and biotic stress including disease and insect pest are the main factors that affect plant growth and limit agricultural productivity. In recent years, with the rapid development of molecular biology, genome editing techniques have been widely used in botany and agronomy due to their characteristics of high efficiency, controllable and directional editing. Genome editing techniques have great application potential in breeding resistant varieties. These techniques have achieved remarkable results in resistance breeding of important cereal crops (such as maize, rice, wheat, etc.), vegetable and fruit crops. Among them, CRISPR/Cas (clustered regularly interspaced short palindromic repeats/CRISPR-associated) provides a guarantee for the stability of crop yield worldwide. In this paper, the development of CRISRR/Cas and its application in different resistance breeding of important crops are reviewed, the advantages and importance of CRISRR/Cas technology in breeding are emphasized, and the possible problems are pointed out.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2022-09-07","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9524261/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"40393335","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}

{"title":"<i>Agrobacterium</i>- and a single Cas9-sgRNA transcript system-mediated high efficiency gene editing in perennial ryegrass.","authors":"Rahul Kumar,&nbsp;Troy Kamuda,&nbsp;Roshani Budhathoki,&nbsp;Dan Tang,&nbsp;Huseyin Yer,&nbsp;Yunde Zhao,&nbsp;Yi Li","doi":"10.3389/fgeed.2022.960414","DOIUrl":"https://doi.org/10.3389/fgeed.2022.960414","url":null,"abstract":"<p><p>Genome editing technologies provide a powerful tool for genetic improvement of perennial ryegrass, an important forage and turfgrass species worldwide. The sole publication for gene editing in perennial ryegrass used gene-gun for plant transformation and a dual promoter based CRISPR/Cas9 system for editing. However, their editing efficiency was low (5.9% or only one gene-edited plant produced). To test the suitability of the maize Ubiquitin 1 (<i>ZmUbi1</i>) promoter in gene editing of perennial ryegrass, we produced <i>ZmUbi1</i> promoter:<i>RUBY</i> transgenic plants. We observed that <i>ZmUbi1</i> promoter was active in callus tissue prior to shoot regeneration, suggesting that the promoter is suitable for Cas9 and sgRNA expression in perennial ryegrass for high-efficiency production of bi-allelic mutant plants. We then used the <i>ZmUbi1</i> promoter for controlling <i>Cas9</i> and sgRNA expression in perennial ryegrass. A ribozyme cleavage target site between the <i>Cas9</i> and sgRNA sequences allowed production of functional Cas9 mRNA and sgRNA after transcription. Using <i>Agrobacterium</i> for genetic transformation, we observed a 29% efficiency for editing the PHYTOENE DESATURASE gene in perennial ryegrass. DNA sequencing analyses revealed that most <i>pds</i> plants contained bi-allelic mutations. These results demonstrate that the expression of a single Cas9 and sgRNA transcript unit controlled by the <i>ZmUbi1</i> promoter provides a highly efficient system for production of bi-allelic mutants of perennial ryegrass and should also be applicable in other related grass species.</p>","PeriodicalId":73086,"journal":{"name":"Frontiers in genome editing","volume":null,"pages":null},"PeriodicalIF":0.0,"publicationDate":"2022-09-06","publicationTypes":"Journal Article","fieldsOfStudy":null,"isOpenAccess":false,"openAccessPdf":"https://www.ncbi.nlm.nih.gov/pmc/articles/PMC9485938/pdf/","citationCount":null,"resultStr":null,"platform":"Semanticscholar","paperid":"33488702","PeriodicalName":null,"FirstCategoryId":null,"ListUrlMain":null,"RegionNum":0,"RegionCategory":"","ArticlePicture":[],"TitleCN":null,"AbstractTextCN":null,"PMCID":"OA","EPubDate":null,"PubModel":null,"JCR":null,"JCRName":null,"Score":null,"Total":0}