Functional Characterization of the MeSSIII-1 Gene and Its Promoter from Cassava.

Soluble starch synthases (SSs) play important roles in the synthesis of cassava starch. However, the expression characteristics of the cassava SSs genes have not been elucidated. In this study, the MeSSIII-1 gene and its promoter, from SC8 cassava cultivars, were respectively isolated by PCR amplification. MeSSIII-1 protein was localized to the chloroplasts. qRT-PCR analysis revealed that the MeSSIII-1 gene was expressed in almost all tissues tested, and the expression in mature leaves was 18.9 times more than that in tuber roots. MeSSIII-1 expression was induced by methyljasmonate (MeJA), abscisic acid (ABA), and ethylene (ET) hormones in cassava. MeSSIII-1 expression patterns were further confirmed in proMeSSIII-1 transgenic cassava. The promoter deletion analysis showed that the -264 bp to -1 bp MeSSIII-1 promoter has basal activity. The range from -1228 bp to -987 bp and -488 bp to -264 bp significantly enhance promoter activity. The regions from -987 bp to -747 bp and -747 bp to -488 bp have repressive activity. These findings will provide an important reference for research on the potential function and transcriptional regulation mechanisms of the MeSSIII-1 gene and for further in-depth exploration of the regulatory network of its internal functional elements.

Transcriptomic study of the role of MeFtsZ2-1 in pigment accumulation in cassava leaves.

MeFtsZ2-1 is a key gene for plant plastid division, but the mechanism by which MeFtsZ2-1 affects pigment accumulation in cassava (Manihot esculenta Crantz) through plastids remains unclear. We found that MeFtsZ2-1 overexpression in cassava (OE) exhibited darker colors of leaves, with increased levels of anthocyanins and carotenoids. Further observation via Transmission Electron Microscopy (TEM) revealed no apparent defects in chloroplast structure but an increase in the number of plastoglobule in OE leaves. RNA-seq results showed 1582 differentially expressed genes (DEGs) in leaves of OE. KEGG pathway analysis indicated that these DEGs were enriched in pathways related to flavonoid, anthocyanin, and carotenoid biosynthesis. This study reveals the role of MeFtsZ2-1 in cassava pigment accumulation from a physiological and transcriptomic perspective, providing a theoretical basis for improving cassava quality.

Construction of a high-density genetic map based on large-scale marker development in Coix lacryma-jobi L. using specific-locus amplified fragment sequencing (slaf-seq).

Coix lacryma-jobi L. is one of the most economically and medicinally important corns. This study constructed a high-density genetic linkage map of C. lacryma-jobi based on a cross between the parents 'Qianyi No. 2' × 'Wenyi No. 2' and their F2 progeny through high-throughput sequencing and the construction of a specific-locus amplified fragment (SLAF) library. After pre-processing, 325.49 GB of raw data containing 1628 M reads were obtained. A total of 22,944 high-quality SLAFs were identified, among which 3952 SLAFs and 3646 polymorphic markers met the requirements for the construction of a genetic linkage map. The integrated map contained 3605 high-quality SLAFs, which were grouped into ten genetic linkage groups. The total length of the map was 1620.39 cM, with an average distance of 0.45 cM and an average of 360.5 markers per linkage group. This report presents the first high-density genetic map of C. lacryma-jobi. This map was constructed using an F2 population and SLAF-seq approach, which allows the development of a large number of polymorphic markers in a short period. These results provide a platform for precise gene/quantitative trait locus (QTL) mapping, map-based gene separation, and molecular breeding in C. lacryma-jobi. They also help identify a target gene for tracking, splitting quantitative traits, and estimating the phenotypic effects of each QTL for QTL mapping. They are of great significance for improving the efficiency of discovering and utilizing excellent gene resources of C. lacryma-jobi.

Metabolic GWAS-based dissection of genetic basis underlying nutrient quality variation and domestication of cassava storage root.

BACKGROUND: Metabolites play critical roles in regulating nutritional qualities of plants, thereby influencing their consumption and human health. However, the genetic basis underlying the metabolite-based nutrient quality and domestication of root and tuber crops remain largely unknown. RESULTS: We report a comprehensive study combining metabolic and phenotypic genome-wide association studies to dissect the genetic basis of metabolites in the storage root (SR) of cassava. We quantify 2,980 metabolic features in 299 cultivated cassava accessions. We detect 18,218 significant marker-metabolite associations via metabolic genome-wide association mapping and identify 12 candidate genes responsible for the levels of metabolites that are of potential nutritional importance. Me3GT, MeMYB4, and UGT85K4/UGT85K5, which are involved in flavone, anthocyanin, and cyanogenic glucoside metabolism, respectively, are functionally validated through in vitro enzyme assays and in vivo gene silencing analyses. We identify a cluster of cyanogenic glucoside biosynthesis genes, among which CYP79D1, CYP71E7b, and UGT85K5 are highly co-expressed and their allelic combination contributes to low linamarin content. We find MeMYB4 is responsible for variations in cyanidin 3-O-glucoside and delphinidin 3-O-rutinoside contents, thus controlling SR endothelium color. We find human selection affects quercetin 3-O-glucoside content and SR weight per plant. The candidate gene MeFLS1 is subject to selection during cassava domestication, leading to decreased quercetin 3-O-glucoside content and thus increased SR weight per plant. CONCLUSIONS: These findings reveal the genetic basis of cassava SR metabolome variation, establish a linkage between metabolites and agronomic traits, and offer useful resources for genetically improving the nutrition of cassava and other root crops.

Experimental Study on the Backflow Mechanism of Proppants in Induced Fractures and Fiber Sand Control Under the Condition of Large-Scale and Fully Measurable Flow Field.

The proppant backflow in the process of flowback has a great significant effect on gas field development. Therefore, the study of proppant backflow is of great significance for the development and production of gas wells. At present, the physical simulation methods for proppant backflow mainly include the tube perforation model, the slot model, an API standard flow tester, and a large-scale flowback apparatus. The current experimental methods are unable to observe the backflow of proppants during the process of the flowback test. In addition, the only characterization parameter for proppant backflow is the liquid flow rate corresponding to the sand discharge in the diversion chamber called critical velocity, which is too simple and single to accurately characterize the movement state of proppants during the flowback process. In this paper, a physical simulation method of proppant backflow in fractures based on the measurement of flow field was proposed. It can realize the observation and fine description of the proppant backflow state and movement rule. In addition, the process of proppant backflow can be quantitatively described by a multidimensional characterization parameter. The research shows that (1) the proppant backflow is closely related to the shape of the sand bank formed during the proppant placement and the irregular voids formed; (2) the fiber increases the strength of the proppant pack significantly; (3) the critical velocity with fiber increased by 2.25 times compared with the critical velocity without fiber, the optimum fiber concentration was 0.8%, and the fiber length was 12 mm; (4) the full fiber injection was selected as the best injection mode by the experiment; and (5) the whole process of flowback can be divided into two stages. In the strong fluid shear stage, the effect of fiber sand control is more significant. However, when the flowback enters the stage of slow erosion, the difference in the sand control effect under different parameters is no longer significant.

MeGLYI-13, a Glyoxalase I Gene in Cassava, Enhances the Tolerance of Yeast and Arabidopsis to Zinc and Copper Stresses.

Although zinc and copper are the two essential nutrients necessary for plant growth, their excessive accumulation in soil not only causes environmental pollution but also seriously threatens human health and inhibits plant growth. The breeding of plants with novel zinc or copper toxicity tolerance capacities represents one strategy to address this problem. Glyoxalase I (GLYI) family genes have previously been suggested to be involved in the resistance to a wide range of abiotic stresses, including those invoked by heavy metals. Here, a MeGLYI-13 gene cloned from a cassava SC8 cultivar was characterized with regard to its potential ability in resistance to zinc or copper stresses. Sequence alignment indicated that MeGLYI-13 exhibits sequence differences between genotypes. Transient expression analysis revealed the nuclear localization of MeGLYI-13. A nuclear localization signal (NLS) was found in its C-terminal region. There are 12 Zn2+ binding sites and 14 Cu2+ binding sites predicted by the MIB tool, of which six binding sites were shared by Zn2+ and Cu2+. The overexpression of MeGLYI-13 enhanced both the zinc and copper toxicity tolerances of transformed yeast cells and Arabidopsis seedlings. Taken together, our study shows the ability of the MeGLYI-13 gene to resist zinc and copper toxicity, which provides genetic resources for the future breeding of plants resistant to zinc and copper and potentially other heavy metals.

Integrated Analysis of Morphological, Physiological, Anatomical and Molecular Responses of Cassava Seedlings to Different Light Qualities.

Light quality is highly important for growth control of in vitro plant cultures. Here, we investigated the effect of blue light (BL), red light (RL) and combined red and blue light (RBL) on in vitro cassava growth. Our results indicate that RL facilitated radial elongation of cassava and increased stomatal conductance as well as glucose, sucrose, fructose and starch content in leaves and cellulose content in the stem. It also enhanced SOD and POD activities but decreased the stomatal density and chlorophyll and carotenoid content in leaves. In addition, RL leads to shorter palisade cells, denser chloroplasts and more starch granules. These phenotypic changes were inverted following BL treatment. The expression levels of photosynthesis-related genes MeLHCA1, MeLHCA3, MePSB27-2, MePSBY, MePETE1 and MePNSL2 in leaves were at their lowest following RL treatment, while the expression levels of MePSB27-2, MePSBY, MePETE1 and MePNSL2 were at their highest after BL treatment. The phenotypic changes after RBL treatment were between the values observed for the RL and BL treatments alone. Moreover, the responses of SC8 and SC9 cassava varieties to light quality were largely conserved. As such, we believe that the results of this study lay the foundation for controlling the in vitro growth of cassava seedlings by light quality.

Development of cassava common mosaic virus-based vector for protein expression and gene editing in cassava.

BACKGROUND: Plant virus vectors designed for virus-mediated protein overexpression (VOX), virus-induced gene silencing (VIGS), and genome editing (VIGE) provide rapid and cost-effective tools for functional genomics studies, biotechnology applications and genome modification in plants. We previously reported that a cassava common mosaic virus (CsCMV, genus Potexvirus)-based VIGS vector was used for rapid gene function analysis in cassava. However, there are no VOX and VIGE vectors available in cassava. RESULTS: In this study, we developed an efficient VOX vector (CsCMV2-NC) for cassava by modifying the CsCMV-based VIGS vector. Specifically, the length of the duplicated putative subgenomic promoter (SGP1) of the CsCMV CP gene was increased to improve heterologous protein expression in cassava plants. The modified CsCMV2-NC-based VOX vector was engineered to express genes encoding green fluorescent protein (GFP), bacterial phytoene synthase (crtB), and Xanthomonas axonopodis pv. manihotis (Xam) type III effector XopAO1 for viral infection tracking, carotenoid biofortification and Xam virulence effector identification in cassava. In addition, we used CsCMV2-NC to deliver single guide RNAs (gMePDS1/2) targeting two loci of the cassava phytoene desaturase gene (MePDS) in Cas9-overexpressing transgenic cassava lines. The CsCMV-gMePDS1/2 efficiently induced deletion mutations of the targeted MePDS with the albino phenotypes in systemically infected cassava leaves. CONCLUSIONS: Our results provide a useful tool for rapid and efficient heterologous protein expression and guide RNA delivery in cassava. This expands the potential applications of CsCMV-based vector in gene function studies, biotechnology research, and precision breeding for cassava.

Integrated Characterization of Cassava (Manihot esculenta) Pectin Methylesterase (MePME) Genes to Filter Candidate Gene Responses to Multiple Abiotic Stresses.

Plant pectin methylesterases (PMEs) play crucial roles in regulating cell wall modification and response to various stresses. Members of the PME family have been found in several crops, but there is a lack of research into their presence in cassava (Manihot esculent), which is an important crop for world food security. In this research, 89 MePME genes were identified in cassava that were separated into two types (type-Ⅰ and type-Ⅱ) according to the existence or absence of a pro-region (PMEI domain). The MePME gene members were unevenly located on 17 chromosomes, with 19 gene pairs being identified that most likely arose via duplication events. The MePMEs could be divided into ten sub-groups in type-Ⅰ and five sub-groups in type-Ⅱ. The motif analysis revealed 11 conserved motifs in type-Ⅰ and 8 in type-Ⅱ MePMEs. The number of introns in the CDS region of type-Ⅰ MePMEs ranged between one and two, and the number of introns in type-Ⅱ MePMEs ranged between one and nine. There were 21 type-Ⅰ and 31 type-Ⅱ MePMEs that contained signal peptides. Most of the type-Ⅰ MePMEs had two conserved "RK/RLL" and one "FPSWVS" domain between the pro-region and the PME domain. Multiple stress-, hormone- and tissue-specific-related cis-acting regulatory elements were identified in the promoter regions of MePME genes. A total of five co-expressed genes (MePME1, MePME2, MePME27, MePME65 and MePME82) were filtered from different abiotic stresses via the use of UpSet Venn diagrams. The gene expression pattern analysis revealed that the expression of MePME1 was positively correlated with the degree of cassava postharvest physiological deterioration (PPD). The expression of this gene was also significantly upregulated by 7% PEG and 14 °C low-temperature stress, but slightly downregulated by ABA treatment. The tissue-specific expression analysis revealed that MePME1 and MePME65 generally displayed higher expression levels in most tissues than the other co-expressed genes. In this study, we obtain an in-depth understanding of the cassava PME gene family, suggesting that MePME1 could be a candidate gene associated with multiple abiotic tolerance.

Experimental Study on Plugging Behavior of Degradable Diverters in Partially Open Fracture in Temporary Plugging and Diverting Fracturing.

Temporary plugging and diverting fracturing technology (TPDF) has been successfully applied to improve reservoir productivity. In real reservoirs, a considerable number of fractures have relatively rapidly decreasing fracture widths and closed ends. However, the plugging behavior of diverters in this typical fracture called the partially open fracture (POF) is still unclear because of the few related studies. This paper aims to investigate the plugging behavior of diverters at the fracture tip. The 3D-printed fracture model was used to reproduce the partially open fracture, and the morphological characteristics of the partially open fracture and the open fracture were compared based on the scan data. A series of plugging experiments were conducted to monitor the transport behavior of the diverter in partially open fractures through multiple pressure sensors on the fracture model and to investigate the influence of diverter formula and fracture type on plugging behavior. Finally, based on the experimental results, the plugging mechanism of diverters in partially open fractures was analyzed. The plugging experiments show that a higher-pressure distribution appears at the fracture tip when using a combination of fibers and particles, indicating that it is beneficial for the diverter to transport to the tip and form plugging in the fracture, and it should be noted that small changes in particle size and concentration had a significant influence on the plugging performance. Therefore, it is recommended to use a combination of fibers and particles of multiple sizes (maximum particle size not exceeding half of the fracture width) to achieve a better plugging effect. In addition, the plugging behaviors of partially open fractures and open fractures are different. For partially open fractures with widths of 1, 2, and 4 mm, the recommended formula of the diverter is 1 wt % fibers + 1 wt % 0.15 mm particles, 1 wt % fibers + 1 wt % 0.15 mm particles + 1 wt % 1 mm particles, and 1 wt % fibers + 1 wt % 0.15 mm particles + 1 wt % 1 mm particles + 1 wt % 2 mm particles, respectively. The above experimental results provide an experimental and theoretical basis for the application of TPDF in the field.

Cycle kurtosis entropy guided symplectic geometry mode decomposition for detecting faults in rotating machinery.

Strong noise interference or compound fault coupling phenomenon may lead to the failure of fault diagnosis technology. This paper focuses on weak feature extraction and compound faults detection for rotating machinery fault diagnosis and proposes adaptive symplectic geometric mode decomposition (SGMD) using cycle kurtosis entropy. Firstly, an index named cycle kurtosis entropy (CKE) is presented to measure the strength of periodic impulses in a signal. The CKE uses the entropy value of calculating all delay cycle kurtosis (CK) to overcome the shortcomings of the CK in adaptive ability and obtain more stable values. Thirdly, CKE is applied to construct an adaptive slip window with optimal length. This process is called the adaptive window segmentation method, which is mainly used to dig out weak fault features in signals. Finally, CKE is used as the component selection criterion to select the components decomposed by SGMD. The selected components are reconstructed to obtain a de-noised signal. Hilbert envelope analysis is applied to the denoised signal to demodulate the fault characteristic frequency. Numerical simulations and experimental investigations using bearings and gears are performed to testify the property of the presented method. The results indicate that the adaptive slip window can enhance the decomposing ability of SGMD under strong noise condition. Moreover, for the strong periodic impulse identification ability, the cycle kurtosis entropy is suitable to determine the optimal components of SGMD. It is expected that the presented method will be effectively used for fault feature extractions in rotating machinery under stationary running conditions.

Experimental Study on a Liquid-Solid Phase-Change Autogenous Proppant Fracturing Fluid System.

In this paper, a liquid-solid phase-change autogenous proppant fracturing fluid system (LSPCAP) was proposed to solve the problems that was caused by "sand-carrying" in conventional fracturing technology in oil and gas fields. The characteristic of the new fluid system is that no solid particles will be injected in the whole process of fracturing construction except liquids. The fluid itself will transform into solid particles under the formation temperature to resist the closure stress in the fractures. There are two kinds of liquids that make up the new fracturing fluid system. One of the liquids is called phase-change liquid (PCL) which occurs in the liquid-solid phase change under the formation temperature to form solid particles. Another is called nonphase-change liquid (NPCL) which controls the dispersity and size of PCL in the two-phase fluid system. Based on the molecular interaction theory and organic chemistry, bisphenol-A epoxy resin was selected as the building unit of the PCL, and the NPCL consisted of deionized water + nonionic surfactant. The test results indicated that the new fracturing fluid shows the properties of non-Newtonian fluid and has no wall-building property. The new fluid system has good compatibility with the formation fluid, conventional fracturing fluid, and hydrochloric acid. Through the filtration test, the filtration coefficients of PCL, NPCL, and mixture are found to be 1.56 × 10-4 m/s1/2, 2.66 × 10-4 m/s1/2, and 1.7 × 10-4 m/s1/2, respectively, and the damage rate of mixture and NPCL is 18 and 17.7%. The friction test results show that the resistance reduction rate reaches 69% when the volume ratio of PCL and NPCL is 1:10. The shear rate and time only affect the size of the autogenous solid particles, and the sorting coefficient (S) of the particles is 1.04-1.73, indicating good sorting. Crushing resistance and conductivity test results show that the crush rate of autogenous solid particles is 3.56-8.42%. The conductivity of the autogenous solid particles is better than those of quartz sand and ceramsite under a pressure of 10-30 MPa.

Changes in sucrose metabolism patterns affect the early maturation of Cassava sexual tetraploid roots.

BACKGROUND: Cassava (Manihot esculenta Crantz) is an important multiuse crop grown for economic and energy purposes. Its vegetative organs are storage roots, in which the main storage material is starch. The accumulation characteristics of starch in cassava roots can directly affect the yield, starch content and maturation of cassava storage roots. In this study, we used a cassava sexual tetraploid (ST), which showed early maturation heterosis in previous work, as the main test material. We analyzed the sucrose metabolism and starch accumulation characteristics of the ST and its parents from the leaf "source" to the storage root "sink" during different developmental stages and explored the regulatory mechanisms of ST storage root early maturation by combining the transcriptome data of the storage roots during the expansion period. RESULTS: The results showed that the trends in sucrose, glucose and fructose contents in the ST leaves were similar to those of the two parents during different stages of development, but the trends in the ST storage roots were significantly different from those of their parents, which showed high sucrose utilization rates during the early stage of development and decreased utilization capacity in the late developmental stage. Transcriptome data showed that the genes that were expressed differentially between ST and its parents were mainly involved in the degradation and utilization of sucrose in the storage roots, and four key enzyme genes were significantly upregulated (Invertase MeNINV8/MeVINV3, Sucrose synthase MeSuSy2, Hexokinase MeHXK2), while the expressions of key enzyme genes involved in starch synthesis were not significantly different. CONCLUSIONS: The results revealed that the pattern of sucrose degradation and utilization in the cassava ST was different from that of its parents and promoted early maturation in its tuberous roots. Starch accumulation in the ST from sucrose mainly occurred during the early expansion stage of the storage roots, and the starch content during this period was higher than that of both parents, mainly due to the regulation of invertase and hexokinase activities during sucrose metabolism. This study provides a basis for further genetic improvements to cassava traits and for breeding varieties that mature early and are adapted well to provide starch supply requirements.

Distinct common signatures of gut microbiota associated with damp-heat syndrome in patients with different chronic liver diseases.

Background: Chronic hepatitis B (CHB) and non-alcoholic fatty liver disease (NAFLD) are prevalent in China. According to traditional Chinese medicine (TCM) theory, damp-heat (DH) syndrome is common in chronic liver disease. However, the biological characteristics related to quantitative diagnosis remain to be determined. This study aimed to identify the consistent alterations in the gut microbiota associated with DH syndrome in patients with CHB or NAFLD. Methods: A total of 405 individuals were recruited, of which 146 were participants who met the consistent TCM diagnosis by three senior TCM physicians and were typical syndromes. All participants were required to provide fresh stool and serum samples. The gut microbiota was assessed by fecal 16S rRNA gene sequencing, and the serum metabolite profiles of participants were quantified by an ultra-performance liquid chromatography coupled to tandem mass spectrometry (UPLC-MS/MS) system. DH syndrome-related bacteria taxa were identified based on the 146 individuals with typical syndromes and validated in all 405 volunteers. Results: The results showed that CHB and NAFLD patients with typical TCM DH syndrome had consistently elevated serum total bile acid (TBA) levels. Significant alterations in microbial community were observed according to TCM syndromes identification. A total of 870 microbial operational taxonomic units and 21 serum metabolites showed the same variation trends in both the CHB and NAFLD DH syndrome groups. The functional analysis predicts consistent dysregulation of bile acid metabolism. Five genera (Agathobacter, Dorea, Lachnospiraceae_NC2004_group, Subdoligranulum, and unclassified_c__Clostridia) significantly decreased in abundance in patients with DH syndrome. We utilize these five genera combined with TBA to construct a random forest classifier model to predict TCM diagnosis. The diagnostic receiver-operator characteristic (ROC) areas for DH syndrome were 0.818 and 0.791 in internal tenfold cross-validation and the test set based on all 405 individuals, respectively. Conclusion: There are common signatures of gut microbiota associated with DH syndrome in patients with different chronic liver diseases. Serum TBA combined with DH-related genera provides a good diagnostic potential for DH syndrome in chronic liver disease.

Influences of real-time acid-rock reaction heat on etched fracture dimensions during acid fracturing of carbonate reservoirs and field applications.

During acid fracturing, acid-rock reaction heat has a significant influence on temperature profiles in fractures and consequently on etched fracture dimensions, but it is usually neglected or simplified in acid fracturing models. This can lead to misestimating of etched fracture dimensions. A model for calculating real-time acid-rock reaction enthalpy, which is a function of temperature, pressure and volumetric work of carbon dioxide produced by reactions, is coupled into a heat transfer model and a fracture growth model, and its effect on etched fracture dimensions is simulated. True experimental data from SL oilfield in China is used for simulation. The results show that acid-rock reaction heat reduces the effective etched fracture length by around 10%, and the effect of reaction heat on the etched fracture length in limestone is 10%-15% larger than in dolomite. Acid-rock reaction heat makes the etched width profile along a fracture more inhomogeneous. With consideration of acid-rock reaction heat, etched fracture widths are 15%-20% larger near the wellbore and over 20% narrower at fracture tip, and its effects are more intense in limestone than in dolomite. The influences of acid-rock reaction heat on etched fracture dimensions are stronger when the initial formation temperature is lower and when acid of high concentration is used. When the pump rate of acid fracturing is increased, the effect of acid-rock reaction heat on etched fracture dimensions is weakened. The new coupled models were used in carbonate reservoirs in Tarim Basin, China for acid fracturing optimization. A scenario comparison showed that the designed treatment parameters of acid fracturing should be different when acid-rock reaction heat was fully considered. The application of the optimized scenario resulted in at least three folds of production rate increase compared to that before stimulation.

Overexpression of MePMEI1 in Arabidopsis enhances Pb tolerance.

Pb is one of the most ubiquitously distributed heavy metal pollutants in soils and has serious negative effects on plant growth, food safety, and public health. Pectin methylesterase inhibitors (PMEIs) play a pivotal role in regulating the integrity of plant cell walls; however, the molecular basis by which PMEIs promote plant resistance to abiotic stress remains poorly understood. In this study, we identified a novel PMEI gene, MePMEI1, from Manihot esculenta, and determined its role in plant resistance to Pb stress. The expression of MePMEI1 was remarkably upregulated in the roots, stems, and leaves of cassava plants following exposure to Pb stress. An analysis of subcellular localization revealed that the MePMEI1 protein was localized in the cell wall. MePMEI1 inhibited commercial orange peel pectin methyltransferase (PME), and the expression of MePMEI1 in Arabidopsis decreased the PME activity, indicating that MePMEI1 can inhibit PME activity in the cell wall. Additionally, the overexpression of MePMEI1 in Arabidopsis reduced oxidative damage and induced the thickening of cell walls, thus contributing to Pb tolerance. Altogether, the study reports a novel mechanism by which the MePMEI1 gene, which encodes the PMEI protein in cassava, plays an essential role in promoting tolerance to Pb toxicity by regulating the thickness of cell walls. These results provide a theoretical basis for the MePMEI1-mediated plant breeding for increasing heavy metal tolerance and provide insights into controlling Pb pollution in soils through phytoremediation in future studies.

Double/triple hit lymphoma in the gastrointestinal tract: clinicopathological features, PD-L1 expression and screening strategy.

We aimed to detect the clinicopathological features and immune microenvironment of double-hit/triple-hit lymphoma in the gastrointestinal tract (GI-DHL/THL) and identify the best diagnostic strategies. A total of 114 cases, including 15 GI-DHL/THL, 42 non-GI-DHL/THL and 57 control diffuse large B-cell lymphoma (DLBCL) cases, were comparatively analyzed for their clinicopathological characteristics, the expression of the immune-regulatory checkpoint PD-L1 and immune microenvironment. We applied univariate and multivariate analyses to determine predictors of DHL/THL. GI-DHL/THL patients showed a higher prevalence of previous infection with hepatitis B virus (HBV) than those with GI-DLBCL. Morphologically, 87% of cases exhibited features of DLBCL. Regarding immunohistochemistry results, the MYC protein expression and the Ki-67 proliferation index were significantly higher in the GI-DHL/THL group than in the GI-DLBCL group. The main source of PD-L1 expression in DHL was tumor-associated macrophages, whereas some tumor cells were positive for PD-L1 in GI-DLBCL cases, as determined through multiplex immunofluorescence staining. The multivariable logistic analysis suggested that 5 variables, namely, age, Mum1, CD10, MYC, and HBV infection status, reflect the risk of DHL/THL. The GI-DHL/THL group show different clinicopathological features and immune microenvironments from DLBCL, which might suggest that different signaling pathways are involved. More work is needed to elucidate the pathogenic mechanism of GI-DHL/THL.

A Transformation and Genome Editing System for Cassava Cultivar SC8.

Cassava starch is a widely used raw material for industrial production. South Chinese cassava cultivar 8 (Manihot esculenta Crantz cv. SC8) is one of the main locally planted cultivars. In this study, an efficient transformation system for cassava SC8 mediated with Agrobacterium strain LBA4404 was presented for the first time. Cassava friable embryogenic calli (FECs) were transformed through the binary vector pCAMBIA1304 harboring GUS- and GFP-fused genes driven by the CaMV35S promoter. The transformation efficiency was increased in the conditions of Agrobacterium strain cell infection density (OD600 = 0.65), 250 µM acetosyringone induction, and agro-cultivation with wet FECs for 3 days in dark. Based on the optimized transformation protocol, approximately 120-140 independent transgenic lines per mL settled cell volume (SCV) of FECs were created by gene transformation in approximately 5 months, and 45.83% homozygous mono-allelic mutations of the MePDS gene with a YAO promoter-driven CRISPR/Cas9 system were generated. This study will open a more functional avenue for the genetic improvement of cassava SC8.

Ability to Inhibit H+ Transmission of Gemini Surfactants with Different Chain Lengths under Different Ca2+ Circumstances.

To study the ability to inhibit ion transmission of the Gemini surfactant under different Ca2+ circumstances, three kinds of Gemini surfactants with different alkyl chain lengths are synthesized (Cn-4-Cn, n = 12, 14, and 16), which are characterized using 1H NMR, 13C NMR, and Fourier transform infrared spectroscopy. To analyze the property of inhibition of the acid-rock reaction rate, surface tension and contact angle measurements and atomic force microscopy (AFM) results are obtained with different surfactants and under different Ca2+ concentrations. Inhibition rates with different alkyl chain lengths and an acid-etched surface morphology are also studied carefully. The result shows that all cationic Gemini surfactants significantly impact the control of the reaction rate, and the reaction rate decreased remarkably by 44.4% after adding 12-4-12. The ΔG and WA indicate that 12-4-12 has the best adsorption ability on the rock with added Ca2+ compared with the other two Gemini surfactants. It is revealed through the AFM that Ca2+ can significantly change the adsorption morphology of the surfactant. The surfactant adsorption area decreased when Ca2+ is dispersed in the solution as well. These two phenomena can lead to the reduced ability to block H+ of 14-4-14 and 16-4-16. However, the presence of Ca2+ affects the adsorption area of 12-4-12 slightly. Thus, the reaction rate, including that of 12-4-12, is almost unchanged. Because 12-4-12 is adsorbed tightly on the rock surface, H+ can only react with the rock on the unabsorbed dot, resulting in rock surface nonuniformity after being etched, which is beneficial for maintaining the conductivity of the crack.

Genome-Wide Identification of Cassava Glyoxalase I Genes and the Potential Function of MeGLYâ -13 in Iron Toxicity Tolerance.

Glyoxalase I (GLYI) is a key enzyme in the pathway of the glyoxalase system that degrades the toxic substance methylglyoxal, which plays a crucial part in plant growth, development, and stress response. A total of 19 GLYI genes were identified from the cassava genome, which distributed randomly on 11 chromosomes. These genes were named MeGLYI-1-19 and were systematically characterized. Transcriptome data analysis showed that MeGLYIs gene expression is tissue-specific, and MeGLYI-13 is the dominant gene expressed in young tissues, while MeGLYI-19 is the dominant gene expressed in mature tissues and organs. qRT-PCR analysis showed that MeGLYI-13 is upregulated under 2 h excess iron stress, but downregulated under 6, 12, and 20 h iron stress. Overexpression of MeGLYI-13 enhanced the growth ability of transgenic yeast under iron stress. The root growth of transgenic Arabidopsis seedlings was less inhibited by iron toxicity than that of the wild type (WT). Potted transgenic Arabidopsis blossomed and podded under iron stress, but flowering of the WT was significantly delayed. The GLYI activity in transgenic Arabidopsis was improved under both non-iron stress and iron stress conditions compared to the WT. The SOD activity in transgenic plants was increased under iron stress, while the POD and CAT activity and MDA content were decreased compared to that in the WT. These results provide a basis for the selection of candidate genes for iron toxicity tolerance in cassava, and lay a theoretical foundation for further studies on the functions of these MeGLYI genes.