CO Oxidation Promoted by NO Adsorption on RhMn2O3- Cluster Anions.

CO oxidation represents an important model reaction in the gas phase to provide a clear structure-reactivity relationship in related heterogeneous catalysis. Herein, in combination with mass spectrometry experiments and quantum-chemical calculations, we identified that the RhMn2O3- cluster cannot oxidize CO into gas-phase CO2 at room temperature, while the NO preadsorbed products RhMn2O3-[(NO)1,2] are highly reactive in CO oxidation. This discovery is helpful to get a fundamental understanding on the reaction behavior in real-world three-way catalytic conditions where different kinds of reactants coexist. Theoretical calculations were performed to rationalize the crucial roles of preadsorbed NO where the strongly attached NO on the Rh atom can greatly stabilize the products RhMn2O2-[(NO)1,2] during CO oxidation and at the same time works together with the Rh atom to store electrons that stay originally in the attached CO2- unit. The leading result is that the desorption of CO2, which is the rate-determining step of CO oxidation by RhMn2O3-, can be greatly facilitated on the reactions of RhMn2O3-[(NO)1,2] with CO.

Immobilization of a broad host range phage on the peroxidase-like Fe-MOF for colorimetric determination of multiple Salmonella enterica strains in food.

A broad host range phage-based nanozyme (Fe-MOF@SalmpYZU47) was prepared for colorimetric detection of multiple Salmonella enterica strains. The isolation of a broad host range phage (SalmpYZU47) capable of infecting multiple S. enterica strains was achieved. Then, it was directly immobilized onto the Fe-MOF to prepare Fe-MOF@SalmpYZU47, exhibiting peroxidase-like activity. The peroxidase-like activity can be specifically inhibited by multiple S. enterica strains, benefiting from the broad host range capture ability of Fe-MOF@SalmpYZU47. Based on it, a colorimetric detection approach was developed for S. enterica in the range from 1.0 × 102 to 1.0 × 108 CFU mL-1, achieving a low limit of detection (LOD) of 11 CFU mL-1. The Fe-MOF@SalmpYZU47 was utilized for detecting S. enterica in authentic food samples, achieving recoveries ranging from 91.88 to 105.34%. Hence, our proposed broad host range phage-based nanozyme exhibits significant potential for application in the colorimetric detection of pathogenic bacteria.

Physiochemistry and sources of individual particles in response to intensified controls during the 2022 Winter Olympics in Beijing.

To investigate the particle sources before, during, and after the 2022 Beijing Winter Olympic and Paralympic (WOP) in Beijing, ambient particles were passively collected from January to March 2022. The physicochemical properties including morphology, size, shape parameters, and elemental compositions were analyzed by the IntelliSEM EPAS (an advanced computer-controlled scanning electron microscopy [CCSEM] system). Using the user-defined classification rules, 37,174 individual particles were automatically classified into 27 major groups and further attributed to seven major sources based on the source-associated characteristics, including mineral dust, secondary aerosol, combustion/industry, carbonaceous particles, salt-related particles, biological particles, and fiber particles. Our results showed that mineral dust (66.5%), combustion/industry (12.6%), and secondary aerosol (6.3%) were the three major sources in a wide size range of 0.2-42.8 µm. During the Winter Olympic Games period, low emission of anthropogenic particles and favorable meteorological conditions contributed to significantly improved air quality. During the Winter Paralympic Games period, more particles sourced from the dust storm, secondary formed particles, and the adverse meteorological conditions resulted in relatively worse air quality. The secondary aerosol all decreased during the competition period, while increased during the non-competition period. Sulfate-related particles had explosive growth and further aggravate the pollution degree during the non-competition period, especially under adverse meteorological conditions. These results provide microscopic evidence revealing variations of physicochemical properties and sources in response to the control measures and meteorological conditions.

AQP4-A25Q Point Mutation in Mice Depolymerizes Orthogonal Arrays of Particles and Decreases Polarized Expression of AQP4 Protein in Astrocytic Endfeet at the Blood-Brain Barrier.

Aquaporin-4 (AQP4) is characterized by the formation of orthogonal arrays of particles (OAPs) comprising its M1 and M23 isoforms in the plasma membrane. However, the biological importance of OAP formation is obscure. Here, we developed an OAP depolymerization male mouse model by transgenic knock-in of an AQP4-A25Q mutation. Analyses of the mutant brain tissue using blue native polyacrylamide gel electrophoresis, super-resolution imaging, and immunogold electron microscopy revealed remarkably reduced OAP structures and glial endfeet localization of the AQP4-A25Q mutant protein without effects on its overall mRNA and protein expression. AQP4A25Q/A25Q mice showed better survival and neurologic deficit scores when cerebral edema was induced by water intoxication or middle cerebral artery occlusion/reperfusion. The brain water content and swelling of pericapillary astrocytic endfeet processes in AQP4A25Q/A25Q mice were significantly reduced, functionally supporting decreased AQP4 protein expression at the blood-brain barrier. The infarct volume and neuronal damage were also reduced in AQP4A25Q/A25Q mice in the middle cerebral artery occlusion/reperfusion model. Astrocyte activation in the brain was alleviated in AQP4A25Q/A25Q mice, which may be associated with decreased cell swelling. We conclude that the OAP structure of AQP4 plays a key role in its polarized expression in astrocytic endfeet processes at the blood-brain barrier. Therefore, our study provided new insights into intervention of cerebral cellular edema caused by stroke and traumatic brain injury through regulating AQP4 OAP formation.SIGNIFICANCE STATEMENT Aquaporin-4 (AQP4) is characterized by orthogonal arrays of particles (OAPs) comprising the M1 and M23 isoforms in the membrane. Here, an OAP depolymerization male mouse model induced by AQP4-A25Q mutation was first established, and the functions of OAP depolymerization in cerebral edema have been studied. The results revealed that AQP4 lost its OAP structure without affecting AQP4 mRNA and protein levels in AQP4-A25Q mice. AQP4-A25Q mutation mice has neuroprotective effects on cerebral edema induced by water intoxication and middle cerebral artery occlusion/reperfusion through relieving the activation of astrocytes and suppressed microglia-mediated neuroinflammation. We concluded that the OAP structure of AQP4 plays a key role in its polarized expression in astrocytic endfeet processes at the blood-brain barrier. Therefore, our study provided new insights into intervention of cerebral cellular edema caused by stroke and traumatic brain injury through regulating AQP4 OAP formation.

Alexithymia in a Chinese Patient with Chronic Pain and Associated Factors: A Cross-Sectional Study.

BACKGROUND: Alexithymia is more prevalent among those with patients living with chronic pain. Information on the prevalence of alexithymia in Chinese patients with chronic pain and associated factors is limited. AIM: The primary objective of this study was to determine the prevalence of alexithymia, as defined by a score of 61 or greater in the 20-item Toronto Alexithymia Scale (TAS-20), in a Chinese patient with chronic pain. The secondary objective was to investigate the relationship between alexithymia and the clinical and psychological aspects of chronic pain. METHODS: A cross-sectional observational study used the TAS-20 to assess alexithymia of Chinese patients with chronic pain. Sociodemographic and clinical information were obtained and participants filled in the Fear Avoidance Beliefs Questionnaire, Hospital Anxiety and Depression Scale, Pain Catastrophizing Scale, and General Self-efficacy Scale. RESULTS: Of the 346 patients screened, 321 patients living with chronic pain were enrolled into the study. The prevalence of alexithymia among the study population (TAS-20 score ≥61) was 19.6% (95% confidence interval [CI]: 15.3-24.0). The findings showed anxiety (odds ratio [OR] = 2.474; 95% CI, 1.241-4.935), pain catastrophizing (2.649; 1.014-6.921), and self-efficacy (0.952; 0.908-0.988) as independent predictors of alexithymia in patients living with chronic pain. CONCLUSIONS: Patients with chronic pain exhibiting alexithymia were at higher risk of pain catastrophizing, anxiety, and lower self-efficacy, compared with patients without alexithymia. It is important to identify and pay a special attention in clinical practice to patients with chronic pain exhibiting alexithymia, as these individuals are unable to properly express their emotions.

Effectiveness of cognitive behavioral therapy for patients after total knee arthroplasty: a systematic review and meta-analysis.

Total knee arthroplasty (TKA) is considered a common surgical option in patients with end-stage osteoarthritis of the knee. This systematic review and meta-analysis aimed to determine the effectiveness of cognitive behavioral therapy (CBT) for patients after TKA. PubMed, EMBASE, the Cochrane Library, Web of Science and CINAHL were searched for randomized controlled trials (RCTs) from inception to 20 August 2021. Included studies were evaluated with the Cochrane risk-of-bias tool. Six RCTs were included. Our study results demonstrated that a significant reduction in pain catastrophing was seen in patients receiving CBT at post-intervention (SMD -0.48, 95% CI = -0.72 to -0.23, I2 17.2%, p = 0.00) but not in 3-month or 12-month follow-up. There were no significant differences between CBT and usual-care patients regarding pain intensity or knee function at different time-points. This is the first time that meta-analysis has been conducted to determine the effectiveness of CBT for patients after TKA. It is necessary to conduct longer follow-ups, include larger samples and conduct rigorous RCTs to further explore this issue.

Facile CO bond cleavage on polynuclear vanadium nitride clusters V4N5.

Identifying the structural configurations of precursors for CO dissociation is fundamentally interesting and industrially important in the fields of, e.g., Fischer-Tropsch synthesis. Herein, we demonstrated that CO could be dissociated on polynuclear vanadium nitride V4N5- clusters at room temperature, and a key intermediate, with CO in a N-assisted tilted bridge coordination where the C-O bond ruptures easily, was discovered. The reaction was characterized by mass spectrometry, photoelectron spectroscopy, and quantum-chemistry calculations, and the nature of the adsorbed CO on product V4N5CO- was further characterized by a collision-induced dissociation experiment. Theoretical analysis evidences that CO dissociation is predominantly governed by the low-coordinated V and N atoms on the (V3N4)VN- cluster and the V3N4 moiety resembles a support. This finding strongly suggests that a novel mode for facile CO dissociation was identified in a gas-phase cluster study.

Water-Gas Shift Catalyzed by Iridium-Vanadium Oxide Clusters IrVO2- with Iridium in a Rare Oxidation State of -II.

The discovery of compounds containing transition metals with an unusual and well-established oxidation state is vital to enrich our horizon on formal oxidation state. Herein, benefiting from the study of the water-gas shift reaction (CO + H2O → CO2 + H2) mediated with the iridium-vanadium oxide cluster IrVO2-, the missing -II oxidation state of iridium was identified. The reactions were performed by using our newly developed double ion trap reactors that can spatially separate the addition of reactants and are characterized by mass spectrometry and quantum-chemical calculations. This finding makes an important step that all the proposed 13 oxidation states of iridium (+IX to -III) have been known. The iridium atom in the IrVO2- cluster features the Ir═V double bond and resembles chemically the coordinated oxygen atom. A reactivity study demonstrated that the flexible role switch of iridium between an oxygen-atom like (Ir-IIVO2-) and a transition-metal-atom like behavior (Ir+IIVO3-) in different species can drive the water-gas shift reaction in the gas phase under ambient conditions. This result parallels and well rationalizes the extraordinary reactivity of oxide-supported iridium single-atom catalysts in related condensed-phase reactions.

Pain catastrophizing and associated factors in preoperative total knee arthroplasty in Lanzhou, China: a cross-sectional study.

BACKGROUND: Pain catastrophizing in preoperative total knee arthroplasty (TKA) patients is associated with several poorly characterised factors in the literature. This study investigated the current state and associated factors of preoperative pain catastrophizing in patients undergoing TKA. METHODS: This descriptive cross-sectional study was conducted at the orthopedics ward of two tertiary hospitals in Lanzhou, China. Pain catastrophizing was measured using the Chinese versions of the Pain Catastrophizing Scale, Short Form-36 (physical function domain), Numerical Rating Scale, Oxford Knee Score, Hospital Anxiety and Depression Scale, and Life Orientation Test-Revised. RESULTS: The study included 360 participants. Preoperative TKA pain catastrophizing in all patients was high, with a mean score of 24.92 (SD: 12.38). The stepwise multiple linear regression analysis revealed anxiety (ß = 0.548, P < 0.01), education level (ß = - 0.179, P < 0.01), physical function (ß = - 0.156, P < 0.01), and pain intensity during activity (ß = 0.105, P = 0.015) as associated factors for pain catastrophizing, possibly explaining 51.2% of the total variation (F = 95.149, P < 0.01). CONCLUSION: Anxiety was the most relevant factor for pain catastrophizing in patients with preoperative TKA. Lower education levels, poor physical function, and stronger pain intensity during the activity were also associated with pain catastrophizing.

Predictors of Satisfactory Outcomes With Fixed-Bearing Lateral Unicompartmental Knee Arthroplasty: Up to 7-year Follow-Up.

BACKGROUND: There is little literature available examining factors that may predict functional recovery after lateral unicompartmental knee arthroplasty (UKA). The purpose of this study was to report short to mid-term effectiveness and evaluate predictors of better outcome following lateral UKA. METHODS: We retrospectively reviewed 248 patients (260 knees) who underwent lateral UKA from January 2013, with a mean 5-year follow-up. The primary outcome measures comprised the Hospital for Special Surgery (HSS) score and patient satisfaction. Multivariate regression analyses were implemented to investigate associations between these factors with a satisfactory outcome. Implant survival was estimated by Kaplan-Meier analysis. RESULTS: Complete follow-up was available for 186 patients (198 knees). At last follow-up, the HSS scores were changed from 52.1 (range, 38-80) preoperatively to 85.6 (range, 61-98) (P < .001), The OKS improved from 22.8 (range, 16-32) preoperatively to 42.7 (range, 30-47) postoperatively (P < .01). The 5-year survival was 99.5%. The multivariate analysis showed that the following factors tended to obtain a satisfactory outcome: higher proportion of ASA class I (P < .001), diagnosis of primary OA (P = .007), postoperative limb alignment (P = .007), and higher preoperative HSS score (P = .019). Patients with valgus 9°-12° reported the highest HSS scores among different subgroups (P < .001). CONCLUSION: Following lateral UKA, postoperative outcomes were satisfactory in patients with lower ASA scores, diagnosis with primary OA, higher preoperative HSS scores and those with postoperative valgus alignment. It is important to understand these correlations to help appropriate patient selection to obtain optimal function after lateral UKA.

Mindfulness-Based Cognitive Therapy for Treating Chronic Pain A Systematic Review and Meta-analysis.

Chronic pain is a significant public health problem with emotional and disabling factors, which may not completely respond to current medical treatments such as opioids. The systematic review and meta-analysis aimed to examine the effectiveness and safety of MBCT for patients with chronic pain. Database searches of PubMed, Medline, EMBASE, the Cochrane Library, PsycINFO, Web of Science, Scopus and CINAHL up to 15 October 2019. Included studies assessed with the Cochrane risk-of-bias tool. Eight RCTs involved 433 patients, including chronic low back pain, fibromyalgia, migraine, rheumatoid arthritis and mix etiology. MBCT intervention demonstrated a short-term improvement on depression mood [standardized mean difference -0.72; 95% confidence interval = -1.22 to -0.22, p = 0.005] compared with usual care and was associated with short-term improvement in mindfulness compared with non-MBCT [SMD 0.51; 95% CI = 0.01 to 1.01, p = 0.04]. Between-group differences in pain intensity, pain inference and pain acceptance were not significant at short- or long-term follow-up. Compared to active treatments, MBCT intervention not found significant differences in either short- or long-term outcomes. MBCT showed short-term efficacious on depressed mood and mindfulness of chronic pain patients. Longer follow-ups, large sample and rigorous RCTs that can be best understand remaining uncertainties needed.

Assessment of radiolucent lines in patients with lateral unicompartmental knee arthroplasty and the relationship between these lines and the outcome.

PURPOSE: To assess the radiolucent lines (RLLs) around both tibial and femoral components in patients following lateral unicompartmental knee arthroplasty (UKA). METHODS: We performed a retrospective review of the records of a consecutive series of patients who had undergone lateral UKA. The RLLs were assessed with standard anteroposterior and lateral radiographs post-operatively. The patient-reported outcome measures included the Hospital for Special Surgery (HSS) score and Oxford knee score (OKS). The femoral component position (FCP) and femoral-tibial angle (FTA) were also recorded. RESULTS: A total of 198 UKAs that had appropriate radiographs and outcome scores were reviewed with a median follow-up of 33 (range, 12-71) months. The results suggested that 69 cases (34.8%) had RLLs on the standard radiographs. The incidence rates of femoral and tibial physiological RLLs were 11.6% (23/198) and 26% (52/198), respectively, of which 3% (6/198) concerned both components. All RLLs were considered "physiologic lines" that developed within one year after surgery. There were no significant differences among the types of RLLs in any of the outcome measures. No differences in FCP (P = .359) or FTA (P = .111) at the last follow-up were seen. CONCLUSIONS: It was found that one-third of UKAs had RLLs on radiographs following lateral UKA. All RLLs developed within one year after surgery. As a clinical consequence, the development of RLLs does not affect the short-term outcomes after lateral UKA.

MicroRNA-377-3p alleviates IL-1ß-caused chondrocyte apoptosis and cartilage degradation in osteoarthritis in part by downregulating ITGA6.

Increasing evidence indicates that altered expression of microRNAs (miRNAs) is associated with osteoarthritis (OA) progression. In our study, we demonstrated that miR-377-3p is underexpressed in OA-affected cartilage and IL-1ß-treated chondrocytes. Overexpression of miR-377-3p enhanced chondrocyte proliferation and restrained apoptosis and signs of cartilage matrix degradation and of an inflammatory response. Furthermore, ITGA6 was identified as a target gene of miR-377-3p. The latter was found to directly bind to the 3' untranslated region (3'UTR) of ITGA6 mRNA and downregulate ITGA6. In addition, ITGA6 expression was high in OA-affected tissues and negatively correlated with miR-77-3p expression. Overexpression of ITGA6 reversed the effects of miR-377-3p on IL-1ß-caused chondrocyte apoptosis, cartilage matrix degradation, and the inflammatory response. Moreover, bioinformatic analysis and a luciferase assay indicated that miR-377-3p expression is regulated by long noncoding RNA NEAT1, which binds to miR-377-3p and inactivates it. We showed that NEAT1 was highly expressed in OA-affected cartilage, negatively correlated with miR-377-3p levels, and positively correlated with ITGA6 levels. These findings provide information for the development of future treatments of OA, suggesting that miR-377-3p may be a therapeutic target in OA.

Correction to: CRISPR-Cas9 mediated targeted disruption of FAD2-2 microsomal omega-6 desaturase in soybean (Glycine max.L).

An amendment to this paper has been published and can be accessed via the original article.

Catalytic CO Oxidation by O2 Mediated with Single Gold Atom Doped Titanium Oxide Cluster Anions AuTi2 O4-6.

Gas-phase studies on catalytic CO oxidation by O2 mediated with gold-containing heteronuclear metal oxide clusters are vital to obtain the structure-reactivity relationship of supported gold catalysts, while it is challenging to trigger the reactivity of clusters with closed-shell electronic structure in O2 activation. Herein, we identified that CO oxidation by O2 can be catalyzed by the AuTi2 O4-6- clusters, among which AuTi2 O4- with closed-shell electronic structure can effectively activate O2 . The reactions were characterized by mass spectrometry and quantum chemical calculations. Theoretical calculations showed that in the initial stage of O2 activation, the Ti2 O4 moiety in AuTi2 O4- contributes dominantly to activate O2 into superoxide (O2- ⋅) without participation of the Au atom. In subsequent steps, the intimate charge transfer interaction between Au and the Ti2 O4 moiety drives the direct dissociation of the O2- ⋅ unit.

Lateral Unicompartmental knee arthroplasty for a secondary osteonecrosis of the lateral femoral condyle. A case report.

BACKGROUND: Secondary osteonecrosis of the knee is a rare event. There are few reports regarding management of this condition. The aim of the present study is to report treatment outcomes for secondary osteonecrosis of the lateral condyle treated with unicompartmental knee arthroplasty (UKA). CASE PRESENTATION: A 54-year-old woman with idiopathic thrombocytopenic purpura, who received low-dosage corticosteroids, complained of knee pain for 5 years and difficulty walking in the last 5 months. Fixed-bearing lateral UKA was performed under general anesthesia combined with midthigh saphenous nerve block. The patient could walk without ambulation aid shortly after the operation and achieved satisfactory knee joint function at the 6-week follow-up. The knee society score (KSS) increased from 68 to 91. The follow-up period was up to 1 year. There was no pain, loosening, or fracture of the prosthesis at the latest follow-up. CONCLUSIONS: This case study demonstrates successful management of secondary osteonecrosis of the lateral femoral condyle is possible with a fixed bearing lateral UKA. Early diagnosis, rigorous indication, and appropriate surgical techniques were critical to maximizing prosthesis stability in lateral UKA.

Does Unicompartmental Knee Replacement Offer Improved Clinical Advantages Over Total Knee Replacement in the Treatment of Isolated Lateral Osteoarthritis? A Matched Cohort Analysis From an Independent Center.

BACKGROUND: The purpose of this study is to compare the functional and radiographic results, perioperative complications, satisfaction rate, and mid-term survivorship after unicompartmental knee arthroplasty (UKA) and total knee arthroplasty (TKA) for the treatment of lateral compartmental knee osteoarthritis (LCKO). METHODS: Between March 2007 and September 2017, we identified 35 patients with primary TKAs and 121 patients with lateral UKAs (LUKAs) for LCKO with a minimum follow-up of 2 years (mean 5.3 years, range 2-12.4). The matched variables were age, gender, operation side, body mass index, American Society of Anesthesiologist grade, initial diagnosis, osteoarthritis grade in lateral compartment, and follow-up time. All patients were assessed using the Oxford Knee Score, Hospital for Special Surgery score, range of motion, length of hospital stay, satisfaction, and complications. Survivorship of UKA and TKA implants was also compared. RESULTS: At last follow-up, LUKA had a significantly better postoperative Oxford Knee Score, Hospital for Special Surgery score, range of motion, shorter length of hospital time, and higher satisfaction rate than matched TKA group. There were significant differences regarding patellar tendon injury (P = .043), superficial wound infection (P = .028), patellar snapping or impingement (P = .047), and stiffness (P < .001). Five-year survivorships free from revision were similar in both groups (99.2% vs 97.1%, P = .347). CONCLUSION: LUKA for LCKO demonstrated more favorable 5-year results in comparison with TKA. Furthermore, LUKA achieved comparable mid-term survivorship and was less likely to suffer from wound infection and knee stiffness, although not overall surgical complications.

Synthesis of Functionalized Indoles via Palladium-Catalyzed Cyclization of N-(2-allylphenyl) Benzamide: A Method for Synthesis of Indomethacin Precursor.

We developed an efficient method for synthesis of substituted N-benzoylindole via Pd(II)-catalyzed C-H functionalization of substituted N-(2-allylphenyl)benzamide. The reaction showed a broad substrate scope (including N-acetyl and N-Ts substrates) and substituted indoles were obtained in good to excellent yields. The most distinctive feature of this method lies in the high selectivity for N-benzoylindole over benzoxazine, and this is the first example of Pd(II)-catalyzed synthesis of substituted N-benzoylindole. Notably, this new method was applied for the synthesis of key intermediate of indomethacin.

Cause of PM2.5 pollution during the 2016-2017 heating season in Beijing, Tianjin, and Langfang, China.

To investigate the cause of fine particulate matter (particles with an aerodynamic diameter less than 2.5 µm, PM2.5) pollution in the heating season in the North China Plain (specifically Beijing, Tianjin, and Langfang), water-soluble ions and carbonaceous components in PM2.5 were simultaneously measured by online instruments with 1-hr resolution, from November 15, 2016 to March 15, 2017. The results showed extreme severity of PM2.5 pollution on a regional scale. Secondary inorganic ions (SNA, i.e., NO3-+SO42+ NH4+) dominated the water-soluble ions, accounting for 30%-40% of PM2.5, while the total carbon (TC, i.e., OC + EC) contributed to 26.5%-30.1% of PM2.5 in the three cities. SNA were mainly responsible for the increasing PM2.5 pollution compared with organic matter (OM). NO3- was the most abundant species among water-soluble ions, but SO42- played a much more important role in driving the elevated PM2.5 concentrations. The relative humidity (RH) and its precursor SO2 were the key factors affecting the formation of sulfate. Homogeneous reactions dominated the formation of nitrate which was mainly limited by HNO3 in ammonia-rich conditions. Secondary formation and regional transport from the heavily polluted region promoted the growth of PM2.5 concentrations in the formation stage of PM2.5 pollution in Beijing and Langfang. Regional transport or local emissions, along with secondary formation, made great contributions to the PM2.5 pollution in the evolution stage of PM2.5 pollution in Beijing and Langfang. The favourable meteorological conditions and regional transport from a relatively clean region both favored the diffusion of pollutants in all three cities.

CRISPR-Cas9 mediated targeted disruption of FAD2-2 microsomal omega-6 desaturase in soybean (Glycine max.L).

BACKGROUND: Recent innovation in the field of genome engineering encompasses numerous levels of plant genome engineering which attract the substantial excitement of plant biologist worldwide. RNA-guided CRISPR Cas9 system has appeared a promising tool in site-directed mutagenesis due to its innovative utilization in different branches of biology. CRISPR-Cas9 nuclease system have supersedes all previously existed strategies and their associated pitfalls encountered with site-specific mutagenesis. RESULTS: Here we demonstrated an efficient sequence specific integration/mutation of FAD2-2 gene in soybean using CRISPR-Cas9 nuclease system. A single guided RNA sequence was designed with the help of a number of bioinformatics tools aimed to target distinct sites of FAD2-2 loci in soybean. The binary vector (pCas9-AtU6-sgRNA) has been successfully transformed into soybean cotyledon using Agrobacterium tumafacien. Taken together our findings complies soybean transgenic mutants subjected to targeted mutation were surprisingly detected in our target gene. Furthermore, the detection of Cas9 gene, BAR gene, and NOS terminator were carried out respectively. Southern blot analysis confirmed the stable transformation of Cas9 gene into soybean. Real time expression with qRT-PCR and Sanger sequencing analysis confirmed the efficient CRISPR-Cas9/sgRNA induced mutation within the target sequence of FAD2-2 loci. The integration of FAD2-2 target region in the form of substitution, deletions and insertions were achieved with notably high frequency and rare off-target mutagenesis. CONCLUSION: High frequent mutation efficiency was recorded as 21% out of all transgenic soybean plants subjected to targeted mutagenesis. Furthermore, Near-infrared spectroscopy (NIR) indicates the entire fatty acid profiling obtained from the mutants seeds of soybean. A considerable modulation in oleic acid content up to (65.58%) whereas the least level of linoleic acid is (16.08%) were recorded. Based on these finding CRISPR-Cas9 system can possibly sum up recent development and future challenges in producing agronomically important crops.