CrnnCrispr: An Interpretable Deep Learning Method for CRISPR/Cas9 sgRNA On-Target Activity Prediction.

CRISPR/Cas9 is a powerful genome-editing tool in biology, but its wide applications are challenged by a lack of knowledge governing single-guide RNA (sgRNA) activity. Several deep-learning-based methods have been developed for the prediction of on-target activity. However, there is still room for improvement. Here, we proposed a hybrid neural network named CrnnCrispr, which integrates a convolutional neural network and a recurrent neural network for on-target activity prediction. We performed unbiased experiments with four mainstream methods on nine public datasets with varying sample sizes. Additionally, we incorporated a transfer learning strategy to boost the prediction power on small-scale datasets. Our results showed that CrnnCrispr outperformed existing methods in terms of accuracy and generalizability. Finally, we applied a visualization approach to investigate the generalizable nucleotide-position-dependent patterns of sgRNAs for on-target activity, which shows potential in terms of model interpretability and further helps in understanding the principles of sgRNA design.

[Prediction of CRISPR/Cas9 off-target activity using multi-scale convolutional neural network].

Clustered regularly interspaced short palindromic repeat/CRISPR-associated protein 9 (CRISPR/Cas9) is a new generation of gene editing technology, which relies on single guide RNA to identify specific gene sites and guide Cas9 nuclease to edit specific location in the genome. However, the off-target effect of this technology hampers its development. In recent years, several deep learning models have been developed for predicting the CRISPR/Cas9 off-target activity, which contributes to more efficient and safe gene editing and gene therapy. However, the prediction accuracy remains to be improved. In this paper, we proposed a multi-scale convolutional neural network-based method, designated as CnnCRISPR, for CRISPR/Cas9 off-target prediction. First, we used one-hot encoding method to encode the sgRNA-DNA sequence pair, followed by a bitwise or operation on the two binary matrices. Second, the encoded sequence was fed into the Inception-based network for training and evaluating. Third, the well-trained model was applied to evaluate the off-target situation of the sgRNA-DNA sequence pair. Experiments on public datasets showed CnnCRISPR outperforms existing deep learning-based methods, which provides an effective and feasible method for addressing the off-target problems.

Interpretable CRISPR/Cas9 off-target activities with mismatches and indels prediction using BERT.

Off-target effects of CRISPR/Cas9 can lead to suboptimal genome editing outcomes. Numerous deep learning-based approaches have achieved excellent performance for off-target prediction; however, few can predict the off-target activities with both mismatches and indels between single guide RNA (sgRNA) and target DNA sequence pair. In addition, data imbalance is a common pitfall for off-target prediction. Moreover, due to the complexity of genomic contexts, generating an interpretable model also remains challenged. To address these issues, firstly we developed a BERT-based model called CRISPR-BERT for enhancing the prediction of off-target activities with both mismatches and indels. Secondly, we proposed an adaptive batch-wise class balancing strategy to combat the noise exists in imbalanced off-target data. Finally, we applied a visualization approach for investigating the generalizable nucleotide position-dependent patterns of sgRNA-DNA pair for off-target activity. In our comprehensive comparison to existing methods on five mismatches-only datasets and two mismatches-and-indels datasets, CRISPR-BERT achieved the best performance in terms of AUROC and PRAUC. Besides, the visualization analysis demonstrated how implicit knowledge learned by CRISPR-BERT facilitates off-target prediction, which shows potential in model interpretability. Collectively, CRISPR-BERT provides an accurate and interpretable framework for off-target prediction, further contributes to sgRNA optimization in practical use for improved target specificity in CRISPR/Cas9 genome editing. The source code is available at https://github.com/BrokenStringx/CRISPR-BERT.

Benchmarking deep learning methods for predicting CRISPR/Cas9 sgRNA on- and off-target activities.

In silico design of single guide RNA (sgRNA) plays a critical role in clustered regularly interspaced, short palindromic repeats/CRISPR-associated protein 9 (CRISPR/Cas9) system. Continuous efforts are aimed at improving sgRNA design with efficient on-target activity and reduced off-target mutations. In the last 5 years, an increasing number of deep learning-based methods have achieved breakthrough performance in predicting sgRNA on- and off-target activities. Nevertheless, it is worthwhile to systematically evaluate these methods for their predictive abilities. In this review, we conducted a systematic survey on the progress in prediction of on- and off-target editing. We investigated the performances of 10 mainstream deep learning-based on-target predictors using nine public datasets with different sample sizes. We found that in most scenarios, these methods showed superior predictive power on large- and medium-scale datasets than on small-scale datasets. In addition, we performed unbiased experiments to provide in-depth comparison of eight representative approaches for off-target prediction on 12 publicly available datasets with various imbalanced ratios of positive/negative samples. Most methods showed excellent performance on balanced datasets but have much room for improvement on moderate- and severe-imbalanced datasets. This study provides comprehensive perspectives on CRISPR/Cas9 sgRNA on- and off-target activity prediction and improvement for method development.

Discovery of highly potent covalent SARS-CoV-2 3CLpro inhibitors bearing 2-sulfoxyl-1,3,4-oxadiazole scaffold for combating COVID-19.

The coronavirus disease (COVID-19) pandemic, caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2), has emerged as a major public health crisis, posing a significant threat to human well-being. Despite the availability of vaccines, COVID-19 continues to spread owing to the emergence of SARS-CoV-2 mutants. This highlights the urgent need for the discovery of more effective drugs to combat COVID-19. As an important target for COVID-19 treatment, 3C-like protease (3CLpro) plays a crucial role in the replication of SARS-CoV-2. In our previous research, we demonstrated the potent inhibitory activities of compound A1, which contains a 2-sulfonyl-1,3,4-oxadiazole scaffold, against SARS-CoV-2 3CLpro. Herein, we present a detailed investigation of structural optimization of A1 and conduct a study on the structure-activity relationship. Among the various compounds tested, sulfoxide D6 demonstrates a potent irreversible inhibitory activity (IC50 = 0.030 µM) against SARS-CoV-2 3CLpro, as well as a favorable selectivity towards host cysteine proteases such as cathepsin B and cathepsin L. Utilizing mass spectrometry-based peptide profiling, we found that D6 covalently binds to Cys145 of SARS-CoV-2 3CLpro. Some representative compounds, namely C11, D9 and D10 also demonstrates antiviral activity against SARS-CoV-2 in Vero E6 cells. Overall, the investigation of the 2-sulfoxyl-1,3,4-oxadiazole scaffold as a novel cysteine reactive warhead would provide valuable insights into the design of potent covalent 3CLpro inhibitors for COVID-19 treatment.

A practical preparation of bicyclic boronates via metal-free heteroatom-directed alkenyl sp2-C‒H borylation.

Bicyclic boronates play critical roles in the discovery of functional materials and antibacterial agents, especially against deadly bacterial pathogens. Their practical and convenient preparation is in high demand but with great challenge. Herein, we report an efficient strategy for the preparation of bicyclic boronates through metal-free heteroatom-directed alkenyl sp2-C‒H borylation. This synthetic approach exhibits good functional group compatibility, and the corresponding boronates bearing halides, aryls, acyclic and cyclic frameworks are obtained with high yields (43 examples, up to 95% yield). Furthermore, a gram-scale experiment is conducted, and downstream transformations of the bicyclic boronates are pursued to afford natural products, drug scaffolds, and chiral hemiboronic acid catalysts.

Nature-inspired catalytic asymmetric rearrangement of cyclopropylcarbinyl cation.

In nature, cyclopropylcarbinyl cation is often involved in cationic cascade reactions catalyzed by natural enzymes to produce a great number of structurally diverse natural substances. However, mimicking this natural process with artificial organic catalysts remains a daunting challenge in synthetic chemistry. We report a small molecule-catalyzed asymmetric rearrangement of cyclopropylcarbinyl cations, leading to a series of chiral homoallylic sulfide products with good to excellent yields and enantioselectivities (up to 99% enantiomeric excess). In the presence of a chiral SPINOL-derived N-triflyl phosphoramide catalyst, the dehydration of prochiral cyclopropylcarbinols occurs rapidly to generate symmetrical cyclopropylcarbinyl cations, which are subsequently trapped by thione-containing nucleophiles. A subgram-scale experiment and multiple downstream transformations of the sulfide products are further pursued to demonstrate the synthetic utility. Notably, a few heteroaromatic sulfone derivatives could serve as "covalent warhead" in the enzymatic inhibition of severe acute respiratory syndrome coronavirus 2 main protease.

Expression of miRNA-1-3p and its target gene in hair follicle cycle development of Liaoning Cashmere goat.

MicroRNA exerts an important regulatory role in almost all the biological process, including hair follicle development in Liaoning Cashmere goat. In order to improve the Cashmere performance of goat, the regulatory role of microRNA in hair follicle cycle has drawn hotspot attention. However, the molecular mechanisms of miRNA-1-3p involved in hair follicle development are poorly understood. In this study, we found that miRNA-1-3p was less expressed in anagen stage of hair follicle cycle of Cashmere goat than that in telogen stage by using RT-qPCR and immunoblotting analysis, in contrast to the expression pattern of FGF14. The Dual-Luciferase reporter assay was employed to verify the relationship between miRNA-1-3p and FGF14. The results showed that miRNA-1-3p specifically binds to the 3'UTR of FGF14 mRNA, and FGF14 is the target gene of miR-1-3p. In conclusion, this study shows that miRNA-1-3p may regulate hair follicle development in Liaoning Cashmere goats by targeting FGF14.

Description of two novel anaerobic members in the family Clostridiaceae, Anaeromonas gelatinilytica gen.nov., sp. nov., and Anaeromonas frigoriresistens sp. nov., isolated from saline lake sediment.

Cells of members of the family Clostridiaceae, phylum Firmicutes, are generally obligate anaerobic rods. Strains D2Q-14T and D2Q-11T were isolated from sediment of the saline lake Manisi in the Xinjiang Uygur Autonomous Region, PR China. In this study, we employed a polyphasic approach and whole genome analysis of the two isolates. Cells of both isolates were Gram-stain-positive rods that were motile by means of flagella and could utilize sulphate, thiosulphate, elemental sulphur and nitrate as electron acceptors. Phylogenetic analyses based on 16S rRNA gene and whole genome sequences indicated that strains D2Q-14T and D2Q-11T constituted a coherent cluster affiliated to the family Clostridiaceae. In addition, genome analysis revealed that strain D2Q-14Tharboured one nonribosomal peptide synthetase gene cluster, making up 1.4 % of the entire genome. The genome-based analysis, including average nucleotide identity, average amino acid identity and in silico DNA-DNA hybridization, biochemical, phenotypic and chemotaxonomic characterization, indicated that strains D2Q-14T and D2Q-11T represented two novel species of a novel genus in the family Clostridiaceae, for which we propose the names Anaeromonas gelatinilytica gen. nov., sp. nov. and Anaeromonas frigoriresistens sp. nov., with the type strains D2Q-14T (=KCTC 15986T=MCCC 1K04634T) and D2Q-11T (=KCTC 15985T=MCCC 1K04391T), respectively.

SAAED: Embedding and Deep Learning Enhance Accurate Prediction of Association Between circRNA and Disease.

Emerging evidence indicates that circRNA can regulate various diseases. However, the mechanisms of circRNA in these diseases have not been fully understood. Therefore, detecting potential circRNA-disease associations has far-reaching significance for pathological development and treatment of these diseases. In recent years, deep learning models are used in association analysis of circRNA-disease, but a lack of circRNA-disease association data limits further improvement. Therefore, there is an urgent need to mine more semantic information from data. In this paper, we propose a novel method called Semantic Association Analysis by Embedding and Deep learning (SAAED), which consists of two parts, a neural network embedding model called Entity Relation Network (ERN) and a Pseudo-Siamese network (PSN) for analysis. ERN can fuse multiple sources of data and express the information with low-dimensional embedding vectors. PSN can extract the feature between circRNA and disease for the association analysis. CircRNA-disease, circRNA-miRNA, disease-gene, disease-miRNA, disease-lncRNA, and disease-drug association information are used in this paper. More association data can be introduced for analysis without restriction. Based on the CircR2Disease benchmark dataset for evaluation, a fivefold cross-validation experiment showed an AUC of 98.92%, an accuracy of 95.39%, and a sensitivity of 93.06%. Compared with other state-of-the-art models, SAAED achieves the best overall performance. SAAED can expand the expression of the biological related information and is an efficient method for predicting potential circRNA-disease association.

Machine learning-aided risk prediction for metabolic syndrome based on 3 years study.

Metabolic syndrome (MetS) is a group of physiological states of metabolic disorders, which may increase the risk of diabetes, cardiovascular and other diseases. Therefore, it is of great significance to predict the onset of MetS and the corresponding risk factors. In this study, we investigate the risk prediction for MetS using a data set of 67,730 samples with physical examination records of three consecutive years provided by the Department of Health Management, Nanfang Hospital, Southern Medical University, P.R. China. Specifically, the prediction for MetS takes the numerical features of examination records as well as the differential features by using the examination records over the past two consecutive years, namely, the differential numerical feature (DNF) and the differential state feature (DSF), and the risk factors of the above features w.r.t different ages and genders are statistically analyzed. From numerical results, it is shown that the proposed DSF in addition to the numerical feature of examination records, significantly contributes to the risk prediction of MetS. Additionally, the proposed scheme, by using the proposed features, yields a superior performance to the state-of-the-art MetS prediction model, which provides the potential of effective prescreening the occurrence of MetS.

Halorubrum salipaludis sp. nov., isolated from the saline-alkaline soil.

A novel extremely halophilic archaeon, designated WN019T, was isolated from the natural saline-alkali wetland soil of Binhai new district, Tianjin, China. Cells of WN019T were aerobic, motile, and pleomorphic rod-shaped, 0.5-0.8 µm in width and 2.0-2.5 µm in length, and the growth occurred optimally at 33-37 °C, pH 7.5-8.0, and in the presence of 15.0-20.0% (w/v) NaCl. Phylogenetic analyses based on 16S rRNA gene sequence comparison showed that the isolate belonged to the genus Halorubrum and exhibited moderate sequence similarity of 97.8% to Halorubrum saccharovorum JCM 8865T. The major respiratory quinones of strain WN019T were MK-8 and MK-8 (H2), and the major polar lipids were glycolipid (GL), phospholipid (PL), phosphatidylglycerol-sulphate (PGS), phosphatidylglycerol (PG) and phosphatidylglycerol-phosphate-methyl ester (Me-PGP). The DNA G + C content of the strain was 67.4 mol%. The average nucleotide identity (ANI) and digital DNA-DNA hybridization (dDDH) value based on whole genome sequences of strain WN019T and Halorubrum saccharovorum JCM 8865T were 87.5% and 35.4%, respectively. Phenotypic, chemotaxonomic, phylogenetic, and genomic analyses suggested that strain WN019T represents a novel species of the genus Halorubrum, for which the name Halorubrum salipaludis sp. nov. is proposed. The type strain is WN019T (= KCTC 4269T = ACCC 19977T).

Halomonas salipaludis sp. nov., isolated from a saline-alkali wetland soil.

Strain WRN001T, a Gram-staining-negative, strictly aerobic, non-motile bacterium was isolated from the natural saline-alkali wetland soil of Binhai new district, Tianjin, China (38°46' N, 117°13' E). Cells of strain WRN001T were 0.3-0.5 µm in width and 1.5-2.5 µm in length, and the growth occurred optimally at 33-37 °C, pH 7.5-8.0, and in the presence of 8-10% (w/v) NaCl. Based on 16S rRNA gene sequence analysis, the isolate could be affiliated to the genus Halomonas, and the highest 16S rRNA gene sequence similarity of strain WRN001T to its closest relative Halomonas qiaohouensis DSM 26770 T was 97.5%. The size of the genome as presented here was 5,475,884 bp with a G + C content of 63.8 mol %. The major respiratory quinone of strainWRN001T was Q-9, and the dominant fatty acids were summed feature 8, summed feature 3, C10:0, C12:0, C12:0 3-OH, C16:0, and C17:0 cyclo. The major polar lipids were diphosphatidylglycerol (DPG), phosphatidylethanolamine (PE), phosphatidylglycerol (PG), phophatidylcholine (PC), two phospholipids (PL), aminolipid (AL), and three unidentified lipids (L). These data combined with the low digital DDH values between strain WRN001T and the close relative, Halomonas alkalitolerans CGMCC 1.9129 T (42.2%) and based on comparisons with currently available genomes, the highest average nucleotide identity (ANIm) value was 91.4% to Halomonas alkalitolerans CGMCC 1.9129 T (GenBank accession No. GCA_001971685.1). Therefore, we propose a novel species in the genus Halomonas to accommodate this novel isolate: Halomonas salipaludis sp. nov. (type strain WRN001T = KCTC 52853 T = ACCC 19974 T).

Pontibacter flavimaris sp. nov., of the family Hymenobacteraceae, isolated from marine sediment.

A Gram-stain-negative, ovoid or rod-shaped, non-flagellated, motile-by-gliding and aerobic bacteria, designated S10-8T, was isolated from marine sediment of the Yellow Sea. Colonies of strain S10-8T had a pink-red pigmentation and its cells were approximately 0.5-0.8 µm×1.0-2.5 µm in size. Growth occurred at 10-45 °C (optimally at 33-37 °C), in the presence of 0-12.0 % NaCl (optimally at 2.0-5.0 %, w/v) and at pH 5.0-8.5 (optimally at pH 7.0-7.5). Phylogenetic analysis of the 16S rRNA gene indicated that strain S10-8T is a member of the genus Pontibacter within the family Hymenobacteraceae, and the 16S rRNA gene sequence similarity of strain S10-8T to its closest relative Pontibacter actiniarum KCTC 12367T was 96.9 %. Strain S10-8T contained MK-7 as the predominant menaquinone and summed feature 4 (iso-C17:1 I and/or anteiso-C17:1 B) and iso-C15:0 as the major fatty acids. The major polar lipids were phosphatidylethanolamine, an unidentified aminophospholipid and an unidentified lipid. The size of the draft genome was 4 623 791 bp and the G+C content was 53.5 mol%. There were low DNA-DNA hybridization values (<48.3±5.2 %) and average nucleotide identity values (<86.5 %) between strain S10-8T and the most closely related recognized Pontibacter species. Therefore, we propose a novel species in the genus Pontibacter to accommodate the novel isolate: Pontibacter flavimaris sp. nov. (type strain S10-8T=KCTC 42769T=ACCC 19859T).

Catalyst-Controlled Diastereoselectivity Switch in the Asymmetric [3 + 2] Annulation of Isatin-Derived MBH Carbonates and 5-Alkenylthiazol-4(5H)-ones.

Exploration of the diastereodivergent synthesis of spirocyclic oxindoles has been challenging. Herein we report asymmetric [3 + 2] annulations of isatin-derived Morita-Baylis-Hillman (MBH) carbonates and 5-alkenylthiazol-4(5H)-ones. Interestingly, two different chiral catalysts, amide-phosphine and 4-dimethylaminopyridine (DMAP)-thiourea, could lead to two kinds of diastereomeric dispiro oxindoles with three contiguous stereogenic centers. The hexafluoroisopropanol (HFIP) additive plays a vital role in accelerating the reaction and tuning the diastereoselectivity. Moreover, both annulation adducts could be further converted to structurally diverse spirooxindoles.

Detection, Structural Elucidation, and Biological Effects of Diverse N-Acyl-homoserine Lactone Signaling Molecules in the Plant-Promoting Endophytic Bacterium Rhizobium oryzihabitans M15.

Quorum sensing (QS), usually performed by N-acyl-homoserine lactones (AHLs) in Gram-staining-negative bacteria, plays an important role in plant-bacteria interactions. Rhizobium oryzihabitans M15 is a plant-growth-promoting rhizobacterium (PGPR) isolated from rice roots. In this study, we found a QS system in the endogenous plasmid of R. oryzihabitans M15 and detected the activity of AHLs by a bioassay method. We identified five AHL analogues in R. oryzihabitans M15 using liquid chromatography-tandem mass spectrometry (LC-MS). The most dominant AHL analogue was N-(3R-hydroxy-7-cis-tetradecanoyl)-l-homoserine lactone according to nuclear magnetic resonance (NMR) and Mosher's reactions. Furthermore, the rosI mutant abolished AHL production and significantly decreased growth, exopolysaccharide (EPS) production, biofilm formation, and motility compared to the wild-type strain. These results lay the foundation for further investigating the QS regulation mechanism and signal pathway of R. oryzihabitans M15 and its interactions with the host plant.

Salimicrobium humidisoli sp. nov., Isolated from Saline-Alkaline Soil.

A Gram-staining-positive, aerobic, non-endospore-forming, coccus-shaped, non-flagellated, and non-motile bacterium, designated WN024T, was isolated from the natural saline-alkali wetland soil of Binhai new district, Tianjin, China. Cells of strain WN024T were catalase- and oxidase-positive. The isolate was able to grow between 20 and 45 °C (optimal at 33-37 °C), pH 6.5-11.0 (optimal 7.5-8.0), and in the presence of 5.0-25.0% NaCl (optimal at 10.0-15.0%, w/v). The isolate could be affiliated to the genus Salimicrobium and the highest 16S rRNA gene sequence similarity of strain WN024T to its closest relative Salimicrobium salexigens DSM 22782T was 97.9%. The size of the genome was 2,622,223 bp in size with a G + C content of 47.1%. The sole respiratory quinone of strain WN024T was MK-7, the predominant fatty acids were iso-C15:0, anteiso-C15:0 and anteiso-C17:0. The major polar lipids were phosphatidylglycerol (PG), glycolipid (GL), phospholipid (PL) and diphosphatidylglycerol (DPG). The cell-wall diamino acid was meso-diaminopimelic acid. The DNA-DNA hybridization values between strain WN024T and the closest relative S. salexigens DSM 22782T was 47.3 ± 2.3%. The highest average nucleotide identity (ANI) value was 92.3% to S. salexigens DSM 22782T (GenBank Accession No. GCA_900156705.1). Therefore, we propose a novel species in the genus Salimicrobium to accommodate this novel isolate, named Salimicrobium humidisoli sp. nov. The type strain is WN024T (= ACCC 19979T = KCTC 33897T).

Prediction of CRISPR/Cas9 single guide RNA cleavage efficiency and specificity by attention-based convolutional neural networks.

CRISPR/Cas9 is a preferred genome editing tool and has been widely adapted to ranges of disciplines, from molecular biology to gene therapy. A key prerequisite for the success of CRISPR/Cas9 is its capacity to distinguish between single guide RNAs (sgRNAs) on target and homologous off-target sites. Thus, optimized design of sgRNAs by maximizing their on-target activity and minimizing their potential off-target mutations are crucial concerns for this system. Several deep learning models have been developed for comprehensive understanding of sgRNA cleavage efficacy and specificity. Although the proposed methods yield the performance results by automatically learning a suitable representation from the input data, there is still room for the improvement of accuracy and interpretability. Here, we propose novel interpretable attention-based convolutional neural networks, namely CRISPR-ONT and CRISPR-OFFT, for the prediction of CRISPR/Cas9 sgRNA on- and off-target activities, respectively. Experimental tests on public datasets demonstrate that our models significantly yield satisfactory results in terms of accuracy and interpretability. Our findings contribute to the understanding of how RNA-guide Cas9 nucleases scan the mammalian genome. Data and source codes are available at https://github.com/Peppags/CRISPRont-CRISPRofft.

Bacillus salipaludis sp. nov., isolated from saline-alkaline soil.

A Gram-staining-positive, endospore-forming, aerobic strain, designed WN066T, was isolated from saline-alkali wetland soil of Tianjin, China. Phylogenetic analysis based on 16S rRNA gene sequence indicated WN066T was a member of the genus of Bacillus, and most closely related to Bacillus drentensis DSM 15600T (98.9%), Bacillus cucumis CCM 8651T (98.8%), Bacillus bataviensis DSM 15601T (98.7%) and Bacillus niacini DSM 2923T (98.7%). However, the average nucleotide identity (ANI) and digital DNA-DNA hybridization (DDH) values between strain WN066T and the most closely related species were less than the previously proposed cutoff values for differentiating species within the genus, suggesting that this strain represented a novel Bacillus species. The strain grew at 19-42 °C (optimally 33-37 °C) in the presence of 3-20% (w/v) NaCl (optimally 8-12%(w/v)), and at pH 6.5-11.0 (optimally 7.5-8.5). The major cellular fatty acids were iso-C15:0 (24.8%) and anteiso-C15:0 (38.9%). The predominant polar lipids consisted of diphosphatidylglycerol (DPG), phosphatidylglycerol (PG) and phosphatidylethanolamine (PE). The size of the draft genome was 6,213,503 bp in size and had a G + C content of 38.6 mol %. The peptidoglycan contained meso-diaminopimelic acid as the diagnostic diamino acid. On basis of phenotypic, phylogenetic, chemotaxonomic and genomic features, strain WN066T represented a novel species within the genus Bacillus, for which the name B. salipaludis sp. nov. is proposed. The type strain is WN066T (= KCTC 33953T = ACCC 60085T).

Palladium-catalyzed allene synthesis enabled by ß-hydrogen elimination from sp2-carbon.

The rational design based on a deep understanding of the present reaction mechanism is an important, viable approach to discover new organic transformations. ß-Hydrogen elimination from palladium complexes is a fundamental reaction in palladium catalysis. Normally, the eliminated ß-hydrogen has to be attached to a sp3-carbon. We envision that the hydrogen elimination from sp2-carbon is possible by using thoroughly designed reaction systems, which may offer a new strategy for the preparation of allenes. Here, we describe a palladium-catalyzed cross-coupling of 2,2-diarylvinyl bromides and diazo compounds, where a ß-vinylic hydrogen elimination from allylic palladium intermediate is proposed to be the key step. Both aryl diazo carbonyl compounds and N-tosylhydrazones are competent carbene precursors in this reaction. The reaction mechanism is explored by control experiments, KIE studies and DFT calculations.