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Nucleotide base editors in plants have been limited to conversion of cytosine to thymine. Here, we describe a new plant adenine base editor based on an evolved tRNA adenosine deaminase fused to the nickase CRISPR/Cas9, enabling A•T to G•C conversion at frequencies up to 7.5% in protoplasts and 59.1% in regenerated rice and wheat plants. An endogenous gene is also successfully modified through introducing a gain-of-function point mutation to directly produce an herbicide-tolerant rice plant. With this new adenine base editing system, it is now possible to precisely edit all base pairs, thus expanding the toolset for precise editing in plants.

Editing plant genomes is technically challenging in hard-to-transform plants and usually involves transgenic intermediates, which causes regulatory concerns. Here we report two simple and efficient genome-editing methods in which plants are regenerated from callus cells transiently expressing CRISPR/Cas9 introduced as DNA or RNA. This transient expression-based genome-editing system is highly efficient and specific for producing transgene-free and homozygous wheat mutants in the T0 generation. We demonstrate our protocol to edit genes in hexaploid bread wheat and tetraploid durum wheat, and show that we are able to generate mutants with no detectable transgenes. Our methods may be applicable to other plant species, thus offering the potential to accelerate basic and applied plant genome-engineering research. Plant genome editing typically relies upon transgenic intermediates, which is a concern given the current regulatory requirements concerning GMOs. Here, Zhang et al . describe a method to edit wheat genomes by transiently expressing CRISPR/Cas9 DNA or RNA, and are able to generate mutant plants with no detectable transgenes.

TALEN-induced mutation of all homologous copies of a gene that represses resistance to an important wheat pathogen confers a trait that has eluded plant breeders for decades. Sequence-specific nucleases have been applied to engineer targeted modifications in polyploid genomes 1 , but simultaneous modification of multiple homoeoalleles has not been reported. Here we use transcription activator–like effector nuclease (TALEN) 2 , 3 and clustered, regularly interspaced, short palindromic repeats (CRISPR)-Cas9 (refs. 4 , 5 ) technologies in hexaploid bread wheat to introduce targeted mutations in the three homoeoalleles that encode MILDEW-RESISTANCE LOCUS (MLO) proteins 6 . Genetic redundancy has prevented evaluation of whether mutation of all three MLO alleles in bread wheat might confer resistance to powdery mildew, a trait not found in natural populations 7 . We show that TALEN-induced mutation of all three TaMLO homoeologs in the same plant confers heritable broad-spectrum resistance to powdery mildew. We further use CRISPR-Cas9 technology to generate transgenic wheat plants that carry mutations in the TaMLO-A1 allele. We also demonstrate the feasibility of engineering targeted DNA insertion in bread wheat through nonhomologous end joining of the double-strand breaks caused by TALENs. Our findings provide a methodological framework to improve polyploid crops.

Substantial efforts are being made to optimize the CRISPR/Cas9 system for precision crop breeding. The avoidance of transgene integration and reduction of off-target mutations are the most important targets for optimization. Here, we describe an efficient genome editing method for bread wheat using CRISPR/Cas9 ribonucleoproteins (RNPs). Starting from RNP preparation, the whole protocol takes only seven to nine weeks, with four to five independent mutants produced from 100 immature wheat embryos. Deep sequencing reveals that the chance of off-target mutations in wheat cells is much lower in RNP mediated genome editing than in editing with CRISPR/Cas9 DNA. Consistent with this finding, no off-target mutations are detected in the mutant plants. Because no foreign DNA is used in CRISPR/Cas9 RNP mediated genome editing, the mutants obtained are completely transgene free. This method may be widely applicable for producing genome edited crop plants and has a good prospect of being commercialized. Protocols for crop genome editing would ideally be quick, efficient and specific while avoiding integration of transgenes into the genome of edited plants. Here, Liang et al . show that CRISPR/Cas9 ribonucleoproteins can be used to generate genome edited wheat plants in as little as nine weeks.

Background Long noncoding RNAs (lncRNAs) have been reported to be important regulators in pathogenesis of human cancers, including nasopharyngeal carcinoma (NPC). Here, we mainly aimed to explore the mechanisms of LncRNA-SNHG5/ miR-1179/HMGB3 axis in NPC progression. Methods RT-qPCR and Western blot analysis were employed to detect mRNA and protein expressions. CCK-8, Transwell and dual luciferase reporter assays were applied to investigate functions of LncRNA-SNHG5/miR-1179/HMGB3 axis. Results Upregulation of lncRNA-SNHG5 and downregulation of miR-1179 were identified in NPC, which were associated with adverse clinical outcomes. Functionally, upregulation of lncRNA-SNHG5 and downregulation of miR-1179 accelerated NPC cell proliferation, migration and invasion. Furthermore, lncRNA-SNHG5 acted as a molecular sponge of miR-1179 in NPC. Besides that, upregulation of HMGB3 was found in NPC, and knockdown of HMGB3 restrained NPC progression. Moreover, HMGB3, a target of miR-1179, regulated NPC progression by mediating LncRNA-SNHG5/miR-1179 axis. Conclusion LncRNA SNHG5 serves as a tumor promoter in NPC by sponging miR-1179 and upregulating HMGB3.

Gene expression is regulated by multiple processes, and the translation of mRNAs into proteins is an especially critical step. Upstream open reading frames (uORFs) are widespread cis -elements in eukaryotic genes that usually suppress the translation of downstream primary ORFs (pORFs). Here, we describe a protocol for fine-tuning gene translation in plants by editing endogenous uORFs with the CRISPR–Cas9 system. The method we present readily yields transgene-free uorf mutant offspring. We provide detailed protocols for predicting uORFs and testing their effects on downstream pORFs using a dual-luciferase reporter system, designing and constructing single guide RNA (sgRNA)–Cas9 vectors, identifying transgene-free uorf mutants, and finally comparing the mRNA, protein and phenotypic levels of target genes in uorf mutants and controls. Predicting uORFs and confirming their effects in protoplasts takes only 2–3 weeks, and transgene-free mutants with edited target uORFs controlling different levels of pORF translation can be obtained within 4 months. Unlike previous methods, our strategy achieves fine-tuning of gene translation in transgene-free derivatives, which accelerates the analysis of gene function and the improvement of crop traits. In this protocol, the authors describe a method to fine-tune gene expression in plants by editing endogenous upstream ORFs with the CRISPR–Cas9 system to prevent their inhibition of the translation of primary ORFs. This protocol yields transgene-free uorf mutant offspring.

HighlightsThe bioinspired self-sensing actuated gradient hydrogel was developed by a wettability-based method via precipitation of MoO2 nanosheets.Self-sensing actuated gradient hydrogel combined ultrafast thermo-responsive actuation (21° s–1), exceptional photothermal efficiency (3.7 °C s–1) and high sensing properties (GF = 3.94).The first self-sensing remote interaction system based on gradient hydrogel actuators and robotic hands was constructed.The development of bioinspired gradient hydrogels with self-sensing actuated capabilities for remote interaction with soft-hard robots remains a challenging endeavor. Here, we propose a novel multifunctional self-sensing actuated gradient hydrogel that combines ultrafast actuation and high sensitivity for remote interaction with robotic hand. The gradient network structure, achieved through a wettability difference method involving the rapid precipitation of MoO2 nanosheets, introduces hydrophilic disparities between two sides within hydrogel. This distinctive approach bestows the hydrogel with ultrafast thermo-responsive actuation (21° s−1) and enhanced photothermal efficiency (increase by 3.7 °C s−1 under 808 nm near-infrared). Moreover, the local cross-linking of sodium alginate with Ca2+ endows the hydrogel with programmable deformability and information display capabilities. Additionally, the hydrogel exhibits high sensitivity (gauge factor 3.94 within a wide strain range of 600%), fast response times (140 ms) and good cycling stability. Leveraging these exceptional properties, we incorporate the hydrogel into various soft actuators, including soft gripper, artificial iris, and bioinspired jellyfish, as well as wearable electronics capable of precise human motion and physiological signal detection. Furthermore, through the synergistic combination of remarkable actuation and sensitivity, we realize a self-sensing touch bioinspired tongue. Notably, by employing quantitative analysis of actuation-sensing, we realize remote interaction between soft-hard robot via the Internet of Things. The multifunctional self-sensing actuated gradient hydrogel presented in this study provides a new insight for advanced somatosensory materials, self-feedback intelligent soft robots and human–machine interactions.

Arginine plays an important role in the nitrogen (N) cycle because it has the highest ratio of N to carbon among amino acids. In recent years, there has been increased research interest in improving the N use of plants, reducing the use of N fertilizer, and enhancing the tolerance of plants to N deficiency. Here, the function of arginine in the growth of apple ( Malus hupehensis ) under N deficiency was explored. The application of 100 μmol L –1 arginine was effective for alleviating N-deficiency stress. Exogenous arginine promoted the absorption and use of N, phosphorus (P), and potassium (K) under low N stress. The net photosynthetic rate, maximal photochemical efficiency of photosystem II, and chlorophyll content were higher in treated plants than in control plants. Exogenous arginine affected the content of many metabolites, and the content of many amino acids with important functions was significantly increased, such as glutamate and ornithine, which play an important role in the urea cycle. Half of the metabolites were annotated to specialized metabolic pathways, including the synthesis of phenolic substances, flavonoids, and other substances with antioxidant activity. Our results indicate that arginine promotes the plant photosynthetic capacity and alters amino acid metabolism and some antioxidants including phenolic substances and flavonoids to improve the tolerance of apple to N deficiency, possibly through the improvement of arginine content, and the absorption of mineral.

Transformers in the distribution network must have their carbon emissions analyzed in order to meet the “double carbon” goal. Using an oil-immersed distribution transformer as the research object, this paper develops a “cradle-to-grave” carbon accounting model. A life cycle assessment (LCA)-based methodology is proposed to account for carbon emissions and total energy demand over the full life cycle and further analyze the carbon emissions within each stage. A case study is presented using data from a 200 kVA transformer manufactured by a transformer plant in Xinjiang. According to the data, there are 112.18 t of carbon emissions and 798.21 GJ of total energy consumed. Distribution network transformers have carbon emissions of 282 kg, 782 kg, 122.96 kg, 11,079.64 kg, and −88.6 kg throughout the manufacture and manufacturing stage, transportation stage, construction and installation stage, operation stage, and waste recycling stage, respectively. There are 4.12 GJ, 9.06 GJ, 1.34 GJ, 785.97 GJ, and −0.28 GJ in total energy requirements. According to the study, which covered 99.03% of the entire life cycle, the operation stage had the largest percentage of carbon emissions. In addition to streamlining the production process and using more energy-efficient equipment, the waste recovery stage successfully decreased the environmental impact of carbon emissions. Sensitivity analysis shows that the silicon steel sheet and transformer oil has a significant impact on the carbon emissions of distribution network transformers during the life cycle, and the higher the grade of silicon steel sheet, the lower the carbon emissions, and the synthetic ester transformer oil has the most comprehensive performance and the lowest carbon emissions.