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2.
BMC Plant Biol ; 24(1): 626, 2024 Jul 03.
Artigo em Inglês | MEDLINE | ID: mdl-38961401

RESUMO

BACKGROUND: The calmodulin (CaM) and calmodulin-like (CML) proteins play regulatory roles in plant growth and development, responses to biotic and abiotic stresses, and other biological processes. As a popular fruit and ornamental crop, it is important to explore the regulatory mechanism of flower and fruit development of passion fruit. RESULTS: In this study, 32 PeCaM/PeCML genes were identified from passion fruit genome and were divided into 9 groups based on phylogenetic analysis. The structural analysis, including conserved motifs, gene structure and homologous modeling, illustrates that the PeCaM/PeCML in the same subgroup have relative conserved structural features. Collinearity analysis suggested that the expansion of the CaM/CML gene family likely took place mainly by segmental duplication, and the whole genome replication events were closely related with the rapid expansion of the gene group. PeCaM/PeCMLs were potentially required for different floral tissues development. Significantly, PeCML26 had extremely high expression levels during ovule and fruit development compared with other PeCML genes, suggesting that PeCML26 had potential functions involved in the development of passion fruit flowers and fruits. The co-presence of various cis-elements associated with growth and development, hormone responsiveness, and stress responsiveness in the promoter regions of these PeCaM/PeCMLs might contribute to their diverse regulatory roles. Furthermore, PeCaM/PeCMLs were also induced by various abiotic stresses. This work provides a comprehensive understanding of the CaM/CML gene family and valuable clues for future studies on the function and evolution of CaM/CML genes in passion fruit. CONCLUSION: A total of 32 PeCaM/PeCML genes were divided into 9 groups. The PeCaM/PeCML genes showed differential expression patterns in floral tissues at different development stages. It is worth noting that PeCML26, which is highly homologous to AtCaM2, not only interacts with multiple BBR-BPC TFs, but also has high expression levels during ovule and fruit development, suggesting that PeCML26 had potential functions involved in the development of passion fruit flowers and fruits. This research lays the foundation for future investigations and validation of the potential function of PeCaM/PeCML genes in the growth and development of passion fruit.


Assuntos
Calmodulina , Flores , Frutas , Passiflora , Filogenia , Proteínas de Plantas , Passiflora/genética , Passiflora/crescimento & desenvolvimento , Flores/genética , Flores/crescimento & desenvolvimento , Flores/metabolismo , Frutas/genética , Frutas/crescimento & desenvolvimento , Frutas/metabolismo , Calmodulina/genética , Calmodulina/metabolismo , Proteínas de Plantas/genética , Proteínas de Plantas/metabolismo , Regulação da Expressão Gênica de Plantas , Genoma de Planta , Genes de Plantas , Perfilação da Expressão Gênica
3.
Plant Commun ; : 101040, 2024 Jul 11.
Artigo em Inglês | MEDLINE | ID: mdl-39001607

RESUMO

Understanding the behavior of endogenous proteins is crucial for functional genomics, yet their dynamic characterization in plants presents substantial challenges. While mammalian studies have leveraged in-locus tagging with the luminescent HiBiT peptide and genome editing for rapid quantification of native proteins, this approach remained unexplored in plants. Here, we introduce the in-locus HiBiT tagging of rice proteins and demonstrate its feasibility in plants. We found that although traditional HiBiT blotting works in rice, it failed to detect two of the three tagged proteins, which is attributed to the low luminescence activity in plants. To overcome this limitation, we engaged in extensive optimization, culminating in a new luciferin substrate coupled with a refined reaction protocol that enhanced luminescence by up to 6.9-fold. This innovation led to the development of TagBIT (tagging with HiBiT), a robust method for high-sensitivity protein characterization in plants. Our application of TagBIT to seven rice genes illustrates its versatility on endogenous proteins, enabling antibody-free protein blotting, real-time protein quantification via luminescence, in-situ visualization using a cross-breeding strategy, and effective immunoprecipitation for protein interaction analysis. The heritable nature of this system, confirmed across T1 to T3 generations, positions TagBIT as a powerful tool for protein study in plant biology.

4.
Elife ; 132024 Jul 01.
Artigo em Inglês | MEDLINE | ID: mdl-38949652

RESUMO

Tubulin posttranslational modifications (PTMs) modulate the dynamic properties of microtubules and their interactions with other proteins. However, the effects of tubulin PTMs were often revealed indirectly through the deletion of modifying enzymes or the overexpression of tubulin mutants. In this study, we directly edited the endogenous tubulin loci to install PTM-mimicking or -disabling mutations and studied their effects on microtubule stability, neurite outgrowth, axonal regeneration, cargo transport, and sensory functions in the touch receptor neurons of Caenorhabditis elegans. We found that the status of ß-tubulin S172 phosphorylation and K252 acetylation strongly affected microtubule dynamics, neurite growth, and regeneration, whereas α-tubulin K40 acetylation had little influence. Polyglutamylation and detyrosination in the tubulin C-terminal tail had more subtle effects on microtubule stability likely by modulating the interaction with kinesin-13. Overall, our study systematically assessed and compared several tubulin PTMs for their impacts on neuronal differentiation and regeneration and established an in vivo platform to test the function of tubulin PTMs in neurons.


Assuntos
Caenorhabditis elegans , Microtúbulos , Processamento de Proteína Pós-Traducional , Tubulina (Proteína) , Animais , Tubulina (Proteína)/metabolismo , Tubulina (Proteína)/genética , Caenorhabditis elegans/metabolismo , Caenorhabditis elegans/genética , Microtúbulos/metabolismo , Proteínas de Caenorhabditis elegans/metabolismo , Proteínas de Caenorhabditis elegans/genética , Acetilação , Axônios/metabolismo , Axônios/fisiologia , Fosforilação , Regeneração Nervosa , Cinesinas/metabolismo , Cinesinas/genética
5.
Artigo em Inglês | MEDLINE | ID: mdl-38705364

RESUMO

OBJECTIVE: To assess near-infrared preirradiation effects on postexercise lower-limb muscle damage and function and determine optimal dosage. DATA SOURCES: PubMed, Embase, Cochrane Library, EBSCO, Web of Science, China National Knowledge Infrastructure, and Wanfang Data were systematically searched (2009-2023). STUDY SELECTION: Randomized controlled trials of near-infrared preirradiation on lower-limb muscles after fatigue exercise were incorporated into the meta-analysis. Out of 4550 articles screened, 21 met inclusion criteria. DATA EXTRACTION: The included studies' characteristics were independently extracted by 2 authors, with discrepancies resolved through discussion or by a third author. Quality assessment was performed using the Cochrane risk of bias tool and the Grading of Recommendations, Assessment, Development, and Evaluation System. DATA SYNTHESIS: In 21 studies, near-infrared preirradiation on lower-limb muscles inhibited the decline in peak torque (standardized mean difference [SMD], 1.33; 95% confidence interval [CI], 1.08-1.59; p<.001; increasing 27.97±4.87N·m), reduced blood lactate (SMD, -0.2; 95% CI, -0.37 to -0.03; p=.272; decreasing 0.54±0.42mmol/L), decreased creatine kinase (SMD, -2.11; 95% CI, -2.57 to -1.65; p<.001; decreasing 160.07±27.96U/L), and reduced delayed-onset muscle soreness (SMD, -0.53; 95% CI, -0.81 to 0.24; p<.001). Using a 24-hour cutoff revealed 2 trends: treatment effectiveness depended on power and energy density, with optimal effects at 24.16 J/cm2 and 275 J/cm2 for energy, and 36.81 mW/cm2 and 5495 mW/cm2 for power. Noting that out of 21 studies, 19 are from Brazil, 1 from the United States, and 1 from Australia, and the results exhibit high heterogeneity. CONCLUSIONS: Although we would have preferred a more geographic dispersion of laboratories, our findings indicate that near-infrared preirradiation mitigates peak torque decline in lower-limb muscles. Influenced by energy and power density with a 24-hour threshold, optimal energy and power densities are observed at 24.16 J/cm2, 275 J/cm2, 36.81 mW/cm2, and 5495 mW/cm2, respectively. Laser preirradiation also reduces blood lactate, creatine kinase, and delayed-onset muscle soreness.

6.
New Phytol ; 2024 May 27.
Artigo em Inglês | MEDLINE | ID: mdl-38798233

RESUMO

Gene silencing is crucial in crop breeding for desired trait development. RNA interference (RNAi) has been used widely but is limited by ectopic expression of transgenes and genetic instability. Introducing an upstream start codon (uATG) into the 5'untranslated region (5'UTR) of a target gene may 'silence' the target gene by inhibiting protein translation from the primary start codon (pATG). Here, we report an efficient gene silencing method by introducing a tailor-designed uATG-containing element (ATGE) into the 5'UTR of genes in plants, occupying the original start site to act as a new pATG. Using base editing to introduce new uATGs failed to silence two of the tested three rice genes, indicating complex regulatory mechanisms. Precisely inserting an ATGE adjacent to pATG achieved significant target protein downregulation. Through extensive optimization, we demonstrated this strategy substantially and consistently downregulated target protein expression. By designing a bidirectional multifunctional ATGE4, we enabled tunable knockdown from 19% to 89% and observed expected phenotypes. Introducing ATGE into Waxy, which regulates starch synthesis, generated grains with lower amylose, revealing the value for crop breeding. Together, we have developed a programmable and robust method to knock down gene expression in plants, with potential for biological mechanism exploration and crop enhancement.

7.
Fish Shellfish Immunol ; 149: 109532, 2024 Jun.
Artigo em Inglês | MEDLINE | ID: mdl-38579977

RESUMO

C-type lectins (CTLs) execute critical functions in multiple immune responses of crustaceans as a member of pattern recognition receptors (PRRs) family. In this study, a novel CTL was identified from the exoskeleton of the oriental river prawn Macrobrachium nipponense (MnLec3). The full-length cDNA of MnLec3 was 1150 bp with an open reading frame of 723 bp, encoding 240 amino acids. MnLec3 protein contained a signal peptide and one single carbohydrate-recognition domain (CRD). MnLec3 transcripts were widely distributed at the exoskeleton all over the body. Significant up-regulation of MnLec3 in exoskeleton after Aeromonas hydrophila challenged suggested the involvement of MnLec3 as well as the possible function of the exoskeleton in immune response. In vitro tests with recombinant MnLec3 protein (rMnLec3) manifested that it had polysaccharide binding activity, a wide spectrum of bacterial binding activity and agglutination activity only for tested Gram-negative bacteria (Escherichia coli, Vibrio anguillarum and A. hydrophila). Moreover, rMnLec3 significantly promoted phagocytic ability of hemocytes against A. hydrophila in vivo. What's more, MnLec3 interference remarkably impaired the survivability of the prawns when infected with A. hydrophila. Collectively, these results ascertained that MnLec3 derived from exoskeleton took an essential part in immune defense of the prawns against invading bacteria as a PRR.


Assuntos
Aeromonas hydrophila , Sequência de Aminoácidos , Proteínas de Artrópodes , Regulação da Expressão Gênica , Hemócitos , Imunidade Inata , Lectinas Tipo C , Palaemonidae , Fagocitose , Filogenia , Alinhamento de Sequência , Animais , Palaemonidae/imunologia , Palaemonidae/genética , Lectinas Tipo C/genética , Lectinas Tipo C/imunologia , Lectinas Tipo C/química , Proteínas de Artrópodes/genética , Proteínas de Artrópodes/imunologia , Proteínas de Artrópodes/química , Hemócitos/imunologia , Imunidade Inata/genética , Aeromonas hydrophila/fisiologia , Alinhamento de Sequência/veterinária , Regulação da Expressão Gênica/imunologia , Perfilação da Expressão Gênica/veterinária , Sequência de Bases , Exoesqueleto/imunologia , Exoesqueleto/química
8.
Sports Health ; : 19417381241235147, 2024 Apr 08.
Artigo em Inglês | MEDLINE | ID: mdl-38587041

RESUMO

CONTEXT: Nontraumatic knee conditions are common in clinical practice. Existing pharmaceutical and immobilization approaches provide limited pain relief and functional enhancement. Low-intensity bloodflow restriction training (LI-BFRT) is being investigated as a nonpharmacological alternative; however, its efficacy is uncertain. OBJECTIVE: To assess the effectiveness of LI-BFRT for nontraumatic knee conditions and compare it with high-intensity resistance training (HI-RT) and low-intensity resistance training (LI-RT). DATA SOURCES: PubMed, EBSCO, Science Direct, Cochrane Library, China Knowledge Infrastructure, Wanfang Data, and VIP databases were searched until May 30, 2023. STUDY SELECTION: Original randomized controlled trials involving nontraumatic knee joint conditions with interventions consisting mainly of LI-BFRT, HI-RT, or LI-RT. The results assessed mainly pain and muscle performance. STUDY DESIGN: Systematic review and meta-analysis. LEVEL OF EVIDENCE: Level 1. DATA EXTRACTION: Sample characteristics, study design, country, disease, groups, evaluation time, duration, and outcomes were extracted. RESULTS: A total of 13 randomized controlled trials were included in the systematic review. Compared with pretreatment, LI-BFRT significantly alleviated pain (weighted standardized mean difference [SMD], -1.33; 95% CI, -1.62 to -1.05), with better additional effects on hip muscle training (SMD, -3.14; 95% CI, -4.07 to -2.75). Compared with LI-RT, LI-BFRT significantly relieved pain in male patients (SMD, -1.47; 95% CI, -1.92 to -1.01). LI-BFRT significantly increased quadriceps cross-sectional area (SMD, 0.53; 95% CI, 0.27-0.78), knee extension strength (SMD, 0.84; 95% CI, 0.48-1.2), and leg press strength (SMD, 0.64; 95% CI, 0.34-0.94) compared with pretreatment. Its effects were superior to those of LI-RT and similar to those of HI-RT. However, sex differences in muscle strength improvement were observed. CONCLUSION: In patients with nontraumatic knee joint conditions, LI-BFRT effectively alleviated pain, increased muscle cross-sectional area, and enhanced muscle strength. LI-BFRT showed pain relief comparable with that of LI-RT while surpassing LI-RT in muscle growth and strength improvement.

9.
Sci Total Environ ; 914: 169904, 2024 Mar 01.
Artigo em Inglês | MEDLINE | ID: mdl-38185157

RESUMO

Understory vegetation has an important impact on soil organic carbon (SOC) accumulation. However, little is known about how understory vegetation alters soil microbial community composition and how microbial diversity contributes to SOC chemical composition and persistence during subtropical forest restoration. In this study, removal treatments of an understory fern (Dicranopteris dichotoma) were carried out within pine (Pinus massoniana) plantations restored in different years in subtropical China. Soil microbial community composition and microbial diversity were measured using phospholipid fatty acids (PLFAs) biomarkers and high-throughput sequencing, respectively. The chemical composition of SOC was also measured via solid-state 13C nuclear magnetic resonance (13C NMR). Our results showed that fern removal decreased alkyl C by 4.2 % but increased O-alkyl C by 15.6 % on average, leading to a decline of alkyl C/O-alkyl C ratio, suggesting altered chemical composition of SOC and lowered SOC recalcitrance without fern. Fern removal significantly lowered the fungi-to-bacteria ratio, and it also reduced fungal and bacterial diversity. Partial correlation analysis revealed that soil nitrogen availability was a key factor influencing microbial diversity. Bacterial diversity showed a close relationship with the Alkyl C/O-alkyl C ratio following fern removal. Furthermore, the microbial community structure and bacterial diversity were responsible for 18 % and 55 % of the explained variance in the chemical composition of SOC, respectively. Taken together, these analyses jointly suggest that bacterial diversity exerts a greater role than microbial community structure in supporting SOC persistence during understory fern removal. Our study emphasizes the significance of understory ferns in supporting microbial abundance and diversity as a means of altering SOC persistence during subtropical forest restoration.


Assuntos
Gleiquênias , Pinus , Solo/química , Carbono/análise , Florestas , China , Bactérias , Microbiologia do Solo
11.
Am J Transl Res ; 15(4): 2802-2810, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37193144

RESUMO

BACKGROUND: To investigate the incidence of complications such as radial artery occlusion (RAO) after distal or conventional transradial access in percutaneous coronary interventions, and to compare the advantages and disadvantages of those two approaches. METHODS: In this retrospective study, the data of 110 patients received either distal transradial access (dTRA) (n=56 cases) or conventional transradial access (cTRA) (n=54 cases) in percutaneous coronary interventions were analyzed to compare the incidence of RAO. RESULTS: The incidence of RAO in the dTRA group significantly decreased compared with that in the cTRA group (P<0.05). Univariate analysis indicated that smoking (r=0.064, P=0.011), dTRA (r=0.431, P<0.001), cTRA (r=0.088, P=0.015), radial artery spasm (r=-0.021, P=0.016), and postoperative arterial compression time (r=0.081, P<0.001) were exposure factors for the incidence of RAO. In multivariable analysis, independent risk factors for RAO were postoperative arterial compression time (P=0.038) and dTRA (P<0.001). CONCLUSIONS: dTRA shortened the postoperative arterial compression time and decreased the incidence of RAO compared with conventional transradial approach.

12.
Front Plant Sci ; 14: 1134209, 2023.
Artigo em Inglês | MEDLINE | ID: mdl-37063194

RESUMO

In-locus editing of agronomically-important genes to optimize their spatiotemporal expression is becoming an important breeding approach. Compared to intensive studies on mRNA transcription, manipulating protein translation by genome editing has not been well exploited. Here, we found that precise knock-in of a regulating element into the 5'UTR of a target gene could efficiently increase its protein abundance in rice. We firstly screened a translational enhancer (AMVE) from alfalfa mosaic virus using protoplast-based luciferase assays with an 8.5-folds enhancement. Then the chemically modified donor of AMVE was synthesized and targeted inserted into the 5'UTRs of two genes (WRKY71 and SKC1) using CRISPR/Cas9. Following the in-locus AMVE knock-in, we observed up to a 2.8-fold increase in the amount of WRKY71 protein. Notably, editing of SKC1, a sodium transporter, significantly increased salt tolerance in T2 seedlings, indicating the expected regulation of AMVE knock-in. These data demonstrated the feasibility of such in-locus editing to enhance protein expression, providing a new approach to manipulating protein translation for crop breeding.

13.
BMC Cardiovasc Disord ; 23(1): 220, 2023 04 28.
Artigo em Inglês | MEDLINE | ID: mdl-37118659

RESUMO

BACKGROUND: Acute coronary syndrome(ACS) is the leading cause of mortality and disability worldwide. Immune response has been confirmed to play a vital role in the occurrence and development of ACS. The objective of this prospective, multicenter, observational study is to define immune response and their relationship to the occurrence and progressive of ACS. METHODS: This is a multicenter, prospective, observational longitudinal cohort study. The primary outcome is the incidence of major adverse cardiovascular events (MACE) including in-stent restenosis, severe ventricular arrhythmia, heart failure, recurrent angina pectoris, and sudden cardiac death, and stroke one year later after ACS. Demographic characteristics, clinical data, treatments, and outcomes are collected by local investigators. Furthermore, freshly processed samples will be stained and assessed by flow cytometry. The expression of S100A4, CD47, SIRPα and Tim-3 on monocytes, macrophages and T cells in ACS patients were collected. FOLLOW-UP: during hospitalization, 3, 6 and 12 months after discharge. DISCUSSION: It is expected that this study will reveal the possible targets to improve the prognosis or prevent from occurrence of MACE in ACS patients. Since it's a multicenter study, the enrollment rate of participants will be accelerated and it can ensure that the collected data are more symbolic and improve the richness and credibility of the test basis. ETHICS AND DISSEMINATION: This study has been registered in Chinese Clinical Trial Registry Center. Ethical approval was obtained from the Affiliated Hospital of Guizhou Medical University. The dissemination will occur through the publication of articles in international peer-reviewed journals. TRIAL REGISTRATION: Chinese Clinical Trial Registry: ChiCTR2200066382.


Assuntos
Síndrome Coronariana Aguda , Humanos , Síndrome Coronariana Aguda/diagnóstico , Síndrome Coronariana Aguda/terapia , Síndrome Coronariana Aguda/epidemiologia , Estudos Prospectivos , Prognóstico , Monócitos , Estudos Longitudinais , Linfócitos T , Estudos de Coortes , Macrófagos , Estudos Observacionais como Assunto , Estudos Multicêntricos como Assunto
14.
Sci Total Environ ; 877: 162803, 2023 Jun 15.
Artigo em Inglês | MEDLINE | ID: mdl-36914127

RESUMO

The invasion of Spartina alterniflora has caused severe damage to the coastal wetland ecosystem of the Yellow River Delta, China. Flooding and salinity are key factors influencing the growth and reproduction of S. alterniflora. However, the differences in response of S. alterniflora seedlings and clonal ramets to these factors remain unclear, and it is not known how these differences affect invasion patterns. In this paper, clonal ramets and seedlings were studied separately. Through literature data integration analysis, field investigation, greenhouse experiments, and situational simulation, we demonstrated significant differences in the responses of clonal ramets and seedlings to flooding and salinity changes. Clonal ramets have no theoretical inundation duration threshold with a salinity threshold of 57 ppt (part per thousand); Seedlings have an inundation duration threshold of about 11 h/day and a salinity threshold of 43 ppt. The sensitivity of belowground indicators of two propagules-types to flooding and salinity changes was stronger than that of aboveground indicators, and it is significant for clones (P < 0.05). Clonal ramets have a larger potentially invadable area than seedlings in the Yellow River Delta. However, the actual invasion area of S. alterniflora is often limited by the responses of seedlings to flooding and salinity. In a future sea-level rise scenario, the difference in responses to flooding and salinity will cause S. alterniflora to further compress native species habitats. Our research findings can improve the efficiency and accuracy of S. alterniflora control. Management of hydrological connectivity and strict restrictions on nitrogen input to wetlands, for example, are potential new initiatives to control S. alterniflora invasion.


Assuntos
Ecossistema , Plântula , Salinidade , Rios , Áreas Alagadas , Poaceae/fisiologia , China , Células Clonais , Espécies Introduzidas
15.
Sci Total Environ ; 870: 161934, 2023 Apr 20.
Artigo em Inglês | MEDLINE | ID: mdl-36736396

RESUMO

Microorganisms facilitate the recovery of previously degraded soils, such as degraded lands experiencing vegetation restoration and understory expansion, through vital soil functions like nutrient cycling and decomposing organic matter. Despite the role of microorganisms in recovery, little is known about the effects of the process on microbial diversity and function. Here, we performed an understory fern, Dicranopteris dichotoma (Thunb.) Berhn removal treatments nested within three Masson pine (Pinus massoniana L.) plantations with different restoration years in subtropical China. Three ferns treatments including no ferns cover, with ferns cover, and the ferns removal treatments were established to assess the impact of the ferns on soil microbial diversity and function during revegetation and drivers of observed changes. We combined high-throughput sequencing, network structure modeling, and function prediction of soil bacterial and fungal communities to determine microbial diversity and functions. Our results showed that soil bacterial and fungal diversity increased with restoration time. Understory ferns significantly increased soil microbial diversity in the un-restored land but the effect became smaller in two restored sites. Understory ferns significantly increased the relative abundance of bacterial phyla Proteobacteria and Acidobacteria, but decreased that of Chloroflexi and Firmicutes. Furthermore, the presence of ferns increased the abundance of Basidiomycota, but increased the abundance of Ascomycota. Co-occurrence network analysis revealed that the presence of ferns leads to more complex of bacterial networks with more connections, nodes, average degrees, betweenness, and degrees. The functional predictions indicate that aerobic chemoheterotrophy, chemoheterotrophy, and nitrogen fixation functional groups play key roles in the nutrient cycling of soils with ferns cover. The bacterial and fungal community compositions were strongly affected by revegetation and understory ferns as litter biomass and soil nitrogen were identified as the key environmental factors. Our study highlights the role of understory in facilitating microbial diversity and function recovery during degraded lands restoration.


Assuntos
Gleiquênias , Solo , Solo/química , Microbiologia do Solo , Biomassa , Nitrogênio/metabolismo , Bactérias/metabolismo
16.
Curr Opin Biotechnol ; 79: 102875, 2023 02.
Artigo em Inglês | MEDLINE | ID: mdl-36610369

RESUMO

As a powerful genome editing technology, CRISPR/Cas is revolutionizing both fundamental research and crop breeding, and has now evolved into large-scale editing tools that are efficient, simple, and programmable. With such CRISPR screening technologies, the numbers of genome-edited crops are rapidly increasing. Here, we describe the general workflow of a CRISPR screen in plants, including the selection of appropriate editors, genome-wide guide RNA design, pooled library construction, massive transformation, and high-throughput genotyping. We also discuss applications for the screening of candidate genes, the optimization of spatiotemporal expression, the evolution of protein activities, and the establishment of genome-wide libraries of knockout mutant. After considering the current challenges and limitations, we finally envision a virus-mediated strategy to improve CRISPR screens.


Assuntos
Sistemas CRISPR-Cas , Edição de Genes , Sistemas CRISPR-Cas/genética , Genoma de Planta/genética , Produtos Agrícolas/genética , Melhoramento Vegetal , Plantas Geneticamente Modificadas/genética
17.
J Integr Plant Biol ; 65(3): 646-655, 2023 Mar.
Artigo em Inglês | MEDLINE | ID: mdl-36218268

RESUMO

With the widespread use of clustered regularly interspaced palindromic repeats (CRISPR)/CRISPR-associated nuclease (Cas) technologies in plants, large-scale genome editing is increasingly needed. Here, we developed a geminivirus-mediated surrogate system, called Wheat Dwarf Virus-Gate (WDV-surrogate), to facilitate high-throughput genome editing. WDV-Gate has two parts: one is the recipient callus from a transgenic rice line expressing Cas9 and a mutated hygromycin-resistant gene (HygM) for surrogate selection; the other is a WDV-based construct expressing two single guide RNAs (sgRNAs) targeting HygM and a gene of interest, respectively. We evaluated WDV-Gate on six rice loci by producing a total of 874 T0 plants. Compared with the conventional method, the WDV-Gate system, which was characterized by a transient and high level of sgRNA expression, significantly increased editing frequency (66.8% vs. 90.1%), plantlet regeneration efficiency (2.31-fold increase), and numbers of homozygous-edited plants (36.3% vs. 70.7%). Large-scale editing using pooled sgRNAs targeting the SLR1 gene resulted in a high editing frequency of 94.4%, further demonstrating its feasibility. We also tested WDV-Gate on sequence knock-in for protein tagging. By co-delivering a chemically modified donor DNA with the WDV-Gate plasmid, 3xFLAG peptides were successfully fused to three loci with an efficiency of up to 13%. Thus, by combining transiently expressed sgRNAs and a surrogate selection system, WDV-Gate could be useful for high-throughput gene knock-out and sequence knock-in.


Assuntos
Edição de Genes , Oryza , Edição de Genes/métodos , Sistemas CRISPR-Cas , Oryza/genética , Genoma de Planta , Plantas/genética
18.
Ying Yong Sheng Tai Xue Bao ; 33(5): 1199-1206, 2022 May.
Artigo em Chinês | MEDLINE | ID: mdl-35730077

RESUMO

Different treatments of harvest residues will change the quantity and quality of soil organic matter, with direct or indirect effects on the composition and content of soil nutrient. Nitrogen is one of the most important soil nutrients. However, the response of soil organic nitrogen fractions to different harvest residue treatments is still unclear. In this study, harvest residue treatments, including harvest residue removed, residue retained and residue burnt, were set up after clear-cutting a 50-year-old mature Cunninghamia lanceolata forest in Sanming City, Fujian, China. The H2SO4 hydrolysis method was used to determine soil organic nitrogen fractions and their driving factors in the 0-10 cm and 10-20 cm soil layers after 5 years of harvest residue treatments. The results showed that residue retained treatment significantly enhanced the contents of soil organic nitrogen and its liable fractions. In the 0-10 cm soil layer, soil organic nitrogen content under residue retained treatment (3.36 g·kg-1) was 1.5 and 1.3 times as those of residue removed and residue burnt treatments, respectively. Residue retained treatment had the highest contents of labile nitrogen Ⅰ and Ⅱ fractions. In 10-20 cm soil layer, the contents of soil organic nitrogen and labile nitrogen Ⅱ fraction were also significantly higher in residue retained treatment (2.20, 0.73 g·kg-1) than that in residue removed and residue burnt treatments. The labile nitrogen index Ⅱ in residue retained treatment (33.9%) was significantly higher than in residue burnt treatment (26.1%). The contents of total carbon, dissolved organic carbon, dissolved organic nitrogen, microbial biomass under residue retained treatment were the highest in both soil layers. Compared with residue removed treatment, residue retained treatment significantly enhanced the abundance of soil bacteria (Gram-positive bacteria and Gram-negative bacteria) in 0-10 cm soil layer. In 10-20 cm soil layer, residue retained treatment had the highest content of fungi and the lowest content of actinomycetes. Pearson analysis showed that there were significant positive correlations of labile fractions of soil organic nitrogen with total carbon, dissolved organic carbon, dissolved organic nitrogen, microbial biomass carbon, microbial biomass nitrogen, bacteria (Gram-positive bacteria, Gram-negative bacteria), and fungi, and negative correlations with actinomycetes. It was concluded that the retention of harvest residue was beneficial to increase the content of soil organic nitrogen and labile fractions, improve soil biochemical properties and had a positive effect on soil microbial community composition. Retention of harvest residue was an effective management measure to maintain soil fertility and improve forest productivity.


Assuntos
Actinobacteria , Cunninghamia , Carbono/análise , China , Florestas , Nitrogênio/análise , Solo/química , Microbiologia do Solo
20.
Biomed Res Int ; 2022: 1336184, 2022.
Artigo em Inglês | MEDLINE | ID: mdl-35463986

RESUMO

Objective: By detecting the levels of external counterpulsation combined with exercise therapy on the levels of moesin, angiopoietin-like protein2 (Angptl 2), angiopoietin-like protein (Angptl 3), hypoxia inducible factor-1α (HIF-1α), and RNA-34a (miR-34a) in patients with coronary artery occlusive disease, the effect of external counterpulsation combined with exercise therapy on the establishment of occluded coronary collateral circulation was studied. Methods: A retrospective analysis of 166 patients with coronary heart disease was confirmed by coronary angiography results that at least one coronary artery (anterior descending branch, circumflex branch, and right coronary artery) was completely occluded and was classified into the control group (routine medication) and the treatment group (routine drug therapy plus exercise therapy and external counterpulsation) according to the treatment plan of the patient. The serum levels of moesin, Angptl 2, Angptl 3, and HIF-1α were detected by enzyme-linked immunosorbent assay (ELISA) test. The index of microcirculatory resistance (IMR) and coronary flow reserve (CFR) of the two groups of patients were measured before and after 2 weeks of treatment. The formation of collateral circulation was analyzed according to the Rentrop classification method. Results: After treatment, the IMR levels of the two groups were significantly decreased, and the CFR levels were significantly increased. The decrease of IMR level and the increase of CFR level in the experimental group were better than those in the control group (P < 0.05). There was no significant difference in the positive detection rate of moesin antibody between the two groups, but the OD detection value of the treatment group decreased significantly (P < 0.05). The levels of Angptl 2, Angptl 3, and miR-34a in the treatment group were lower than those in the control group, while the relative expression of HIF-1α was higher than that in the control group. The difference was statistically significant (P < 0.05). External counterpulsation combined with exercise therapy improved the formation rate of collateral circulation (P < 0.05). Conclusions: External counterpulsation combined with exercise therapy can reduce moesin antibody, Angptl 2, Angptl 3, and miR-34a levels increase HIF-1α levels, and promote the establishment of occluded coronary collateral circulation.


Assuntos
Circulação Colateral , Doença da Artéria Coronariana , Contrapulsação , Terapia por Exercício , MicroRNAs , Angiopoietinas , Doença da Artéria Coronariana/fisiopatologia , Doença da Artéria Coronariana/terapia , Circulação Coronária , Vasos Coronários , Contrapulsação/métodos , Humanos , Microcirculação , Estudos Retrospectivos
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