Differential responses of contrasting low phosphorus tolerant cotton genotypes under low phosphorus and drought stress.

BACKGROUND: Drought is one of the main reasons for low phosphorus (P) solubility and availability. AIMS: The use of low P tolerant cotton genotypes might be a possible option to grow in drought conditions. METHODS: This study investigates the tolerance to drought stress in contrasting low P-tolerant cotton genotypes (Jimian169; strong tolerant to low P and DES926; weak tolerant to low P). In hydroponic culture, the drought was artificially induced with 10% PEG in both cotton genotypes followed by low (0.01 mM KH2PO4) and normal (1 mM KH2PO4) P application. RESULTS: The results showed that under low P, PEG-induced drought greatly inhibited growth, dry matter production, photosynthesis, P use efficiency, and led to oxidative stress from excessive malondialdehyde (MDA) and higher accumulation of reactive oxygen species (ROS), and these effects were more in DES926 than Jimian169. Moreover, Jimian169 alleviated oxidative damage by improving the antioxidant system, photosynthetic activities, and an increase in the levels of osmoprotectants like free amino acids, total soluble proteins, total soluble sugars, and proline. CONCLUSIONS: The present study suggests that the low P-tolerant cotton genotype can tolerate drought conditions through high photosynthesis, antioxidant capacity, and osmotic adjustment.

Integrative physiological, transcriptome and metabolome analysis reveals the involvement of carbon and flavonoid biosynthesis in low phosphorus tolerance in cotton.

Phosphorus (P) is an essential nutrient controlling plant growth and development through the regulation of basic metabolic processes; however, the molecular details of these pathways remain largely unknown. In this study, physiological, transcriptome, and metabolome analysis were compared for two cotton genotypes with different low P tolerance under P starvation and resupply. The results showed that the glucose, fructose, sucrose, and starch contents increased by 18.2%, 20.4%, 20.2%, and 14.3% in the roots and 18.3%, 23.3%, 11.0%, and 13.6% in the shoot of Jimian169 than DES926, respectively. Moreover, the activities of enzymes related to carbon and phosphorus metabolism were higher in the roots and shoots of Jimian169 than DES926. In addition, transcriptome analysis revealed that the number of differentially expressed genes (DEGs) was higher in both roots (830) and shoots (730) under P starvation and the DEGs drastically reduced upon P resupply. The KEGG analysis indicated that DEGs were mainly enriched in phenylpropanoid biosynthesis, carbon metabolism, and photosynthesis. The metabolome analysis showed the enrichment of phenylpropanoid, organic acids and derivatives, and lipids in all the pairs at a given time point. The combined transcriptome and metabolome analysis revealed that carbon metabolism and flavonoid biosynthesis are involved in the P starvation response in cotton. Moreover, co-expression network analysis identified 3 hub genes in the roots and shoots that regulate the pathways involved in the P starvation response. This study provides the foundation for understanding the mechanisms of low P tolerance and the hub genes as a potential target for the development of low P tolerant genotypes.

Phosphorus and carbohydrate metabolism contributes to low phosphorus tolerance in cotton.

Low phosphorus (P) is one of the limiting factors in sustainable cotton production. However, little is known about the performance of contrasting low P tolerant cotton genotypes that might be a possible option to grow in low P condition. In the current study, we characterized the response of two cotton genotypes, Jimian169 a strong low P tolerant, and DES926 a weak low P tolerant genotypes under low and normal P conditions. The results showed that low P greatly inhibited growth, dry matter production, photosynthesis, and enzymatic activities related to antioxidant system and carbohydrate metabolism and the inhibition was more in DES926 as compared to Jimian169. In contrast, low P improved root morphology, carbohydrate accumulation, and P metabolism, especially in Jimian169, whereas the opposite responses were observed for DES926. The strong low P tolerance in Jimian169 is linked with a better root system and enhanced P and carbohydrate metabolism, suggesting that Jimian169 is a model genotype for cotton breeding. Results thus indicate that the Jimian169, compared with DES926, tolerates low P by enhancing carbohydrate metabolism and by inducing the activity of several enzymes related to P metabolism. This apparently causes rapid P turnover and enables the Jimian169 to use P more efficiently. Moreover, the transcript level of the key genes could provide useful information to study the molecular mechanism of low P tolerance in cotton.

Genome-wide expression analysis reveals involvement of asparagine synthetase family in cotton development and nitrogen metabolism.

Asparagine synthetase (ASN) is one of the key enzymes of nitrogen (N) metabolism in plants. The product of ASN is asparagine, which is one of the key compounds involved in N transport and storage in plants. Complete genome-wide analysis and classifications of the ASN gene family have recently been reported in different plants. However, little is known about the systematic analysis and expression profiling of ASN proteins in cotton development and N metabolism. Here, various bioinformatics analysis was performed to identify ASN gene family in cotton. In the cotton genome, forty-three proteins were found that determined ASN genes, comprising of 20 genes in Gossypium hirsutum (Gh), 13 genes in Gossypium arboreum, and 10 genes in Gossypium raimondii. The ASN encoded genes unequally distributed on various chromosomes with conserved glutamine amidotransferases and ASN domains. Expression analysis indicated that the majority of GhASNs were upregulated in vegetative and reproductive organs, fiber development, and N metabolism. Overall, the results provide proof of the possible role of the ASN genes in improving cotton growth, fiber development, and especially N metabolism in cotton. The identified hub genes will help to functionally elucidate the ASN genes in cotton development and N metabolism.

Physiological Characteristics of Cotton Subtending Leaf Are Associated With Yield in Contrasting Nitrogen-Efficient Cotton Genotypes.

Nitrogen (N) plays an important role in various plant physiological processes, but studies on the photosynthetic efficiency and enzymatic activities in the cotton subtending leaves and their contribution to yield are still lacking. This study explored the influence of low, moderate, and high N levels on the growth, photosynthesis, carbon (C) and N metabolizing enzymes, and their contribution to yield in CCRI-69 (N-efficient) and XLZ-30 (N-inefficient). The results showed that moderate to high N levels had significantly improved growth, photosynthesis, and sucrose content of CCRI-69 as compared to XLZ-30. The seed cotton yield and lint yield of CCRI-69 were similar under moderate and high N levels but higher than XLZ-30. Similarly, moderate to high N levels improved the C/N metabolizing enzymatic activities in the subtending leaf of CCRI-69 than XLZ-30. A strong correlation was found between subtending leaf N concentration with C/N metabolizing enzymes, photosynthesis, sucrose contents, boll weight, and seed cotton yield of N-efficient cotton genotype. These findings suggest that subtending leaf N concentration regulates the enzymatic activities and has a key role in improving the yield. These parameters may be considered for breeding N-efficient cotton genotypes, which might help to reduce fertilizer loss and improve crop productivity.

A rapid and accurate method for herpesviral gnome editing / 生物工程学报

To rapidly and accurately manipulate genome such as gene deletion, insertion and site mutation, the whole genome of a very virulent strain Md5 of Marek's disease virus (MDV) was inserted into bacterial artificial chromosome (BAC) through homogeneous recombination. The recombinant DNA was electroporated into DH10B competent cells and identified by PCR and restriction fragment length polymorphism analysis. An infectious clone of Md5BAC was obtained following transfection into chicken embryo fibroblast (CEF) cells. Furthermore, a lorf10 deletion mutant was constructed by two step Red-mediated homologous recombination. To confirm the specific role of gene deletion, the lorf10 was reinserted into the original site of MDV genome to make a revertant strain. All the constructs were rescued by transfection into CEF cells, respectively. The successful packaging of recombinant viruses was confirmed by indirect immunofluorescence assay. The results of growth kinetics assay and plaques area measurement showed that the lorf10 is dispensable for MDV propagation in vitro. Overall, this study successfully constructed an infectious BAC clone of MDV and demonstrated its application in genome manipulation; the knowledge gained from our study could be further applied to other hepesviruses.

Growth and nitrogen metabolism are associated with nitrogen-use efficiency in cotton genotypes.

Crops, including cotton, are sensitive to nitrogen (N) and excessive use can lead to an increase in production costs and environmental problems. We hypothesized that the use of cotton genotypes with substantial root systems and high genetic potentials for nitrogen-use efficiency (NUE) would best address these problems. Therefore, the interspecific variations and traits contributing to NUE in six cotton genotypes having contrasting NUEs were studied in response to various nitrate concentrations. Large genotypic variations were observed in morphophysiological and biochemical traits, especially shoot dry weight, root traits, and N-assimilating enzyme levels. The roots of all the cotton genotypes were more sensitive to low-than high-nitrate concentrations, and the genotype CCRI-69 had the largest root system irrespective of the nitrate concentration. The root morphological traits were positively correlated with N-utilization efficiency and were more affected by genotype than nitrate concentration. Conversely, growth and N-assimilating enzyme levels were more affected by nitrate concentration and were positively correlated with N-uptake efficiency. Based on shoot dry weight, CCRI-69 and XLZ-30 were identified as N-efficient and N-inefficient genotypes, respectively, and these results were confirmed by their contrasting root systems, N metabolism, and NUEs. In the future, multi-omics techniques will be performed to identify key genes/pathways involved in N metabolism, which may have the potential to improve root architecture and increase NUE.

Untangling the molecular mechanisms and functions of nitrate to improve nitrogen use efficiency.

A huge amount of nitrogenous fertilizer is used to increase crop production. This leads to an increase in the cost of production, and to human and environmental problems. It is therefore necessary to improve nitrogen use efficiency (NUE) and to design agronomic, biotechnological and breeding strategies for better fertilizer use. Nitrogen use efficiency relies primarily on how plants extract, uptake, transport, assimilate, and remobilize nitrogen. Many plants use nitrate as a preferred nitrogen source. It acts as a signaling molecule in the various important physiological processes required for growth and development. As nitrate is the main source of nitrogen in the soil, root nitrate transporters are important subjects for study. The latest reports have also discussed how nitrate transporter and assimilation genes can be used as molecular tools to improve NUE in crops. The purpose of this review is to describe the mechanisms and functions of nitrate as a specific factor that can be addressed to increase NUE. Improving factors such as nitrate uptake, transport, assimilation, and remobilization through activation by signaling, sensing, and regulatory processes will improve plant growth and NUE. © 2019 Society of Chemical Industry.

Experimental study of HSP70-TKD induced NK cells migrated toward tumor cells / 中国免疫学杂志

Objective:To investigate the Migration ability toward human pancreatic carcinoma cell line and human colon carcinoma cell line with difference HSP 70 plasma membrane expression .Methods: CD3-CD56+NK cells were obtained from human peripheral blood mononuclear(PBMC)in stem cell growth medium SCGM,2μg/ml TKD was added to the medium on 10th day,the ac-tivating receptor CD94/NKG2C expression levels on NK cells was detected with FAC after 4 days.The human pancreatic carcinoma cell line Colo357 and the human colon carcinoma cell line CW 2 were separated into Colo+and CW2+with high HSP70 expression and Colo-and CW2-with low HSP70 expression;Migration assays of NK to the four difference cell lines were performed in a Transwell cell culture system.The cytolytic activity of TKD-activated NK cells against the four subline with HSP 70 expression on their cell surface was analyzed by MTT assay.Results:Flow cytometry analysis showed that CD 3-CD56+NK cells could expanded after 2 weeks in SCGM medium,and the largest percentage of NK cell was (92.50 ±1.25 )%.CD94 expression levels on NK cells increased obviously after TKD inducement the cell surface HSP 70 expression of Colo+, Colo-were ( 78.2 ±2.2 )% and ( 27.3 ±1.2 )% separately , the cell surface HSP70 expression of CW2+,CW2-were (91.1±2.5)%and (18.2±1.0)%separately after FACS;the Migration of NK cells toward Colo+was (68.6±2.8)%,higher than the migration toward Colo-with (22.8±1.5)%;the Migration of NK cells toward CW2+was(73.5±2.7)%,higher than the migration toward CW2-with (18.2±1.3)%;the cytolytic activity of NK against Colo +was(61.2± 3.0)%compared to (24.5 ±1.5)%against Colo-when the ratio of effector cells and target cell was 20 ∶1,the cytolytic activity of NK against CW2+was (63.8±3.2)%compared to (22.4±1.8)% against CW2-when the ratio of effector cells and target cell was 20∶1.Conclusion:TKD-activated NK cells are highly efficient cytolytic effector cells which have stronger significant migration toward HSP70-positive tumor target cells on their cell surface in vitro .

Classification of Bacterial Colonies on Agar Plates Using Hyperspectral Imaging Technology / 分析化学

Rapid detection and classification of bacteria colonies ( Escherichia coli, Listeria monocytogens and Staphylococcus aureus) were investigated by using hyperspectral imaging. The hyperspectral reflectance images (390-1040 nm ) of bacterial colonies on agar plates were collected. Bacterial spectra were extracted automatically based on the masks produced by segmenting a band difference image using the OTSU method. Full wavelength and simplified PLS-DA models were established for classification of bacterial colonies. For the full wavelength model, the overall correct classification rate ( OCCR) and confident OCCR for the prediction set were 100% and 95. 9%, respectively. Besides, competitive adaptive reweighted sampling ( CARS), genetic algorithm ( GA ) and least angle regression-least absolute shrinkage and selection operator ( LARS-Lasso) were used to select feature wavelengths for the development of simplified models. Among them, the CARS-model outperformed the other two in terms of precision, stability and classification accuracy with OCCR and confident OCCR of 100% and 98. 0% for the prediction set, respectively. It was demonstrated that hyperspectral imaging was an effective technology for nondestructive detection of bacterial colonies with high accuracy and high speed. The allocated feature wavelengths by CARS could lay theoretical basis for developing low cost multispectral imaging systems for bacterial colony detection.

Protection of electroacupuncture and compound salviae in coordination on ischemia-reperfusion myocardium / 中国组织工程研究

BACKGROUND: Both electroacupuncture and salviae act on prevention and treatment ofis chemia-reperfusion injury, but, whether such action is achieved or not by coordination of both? OBJECTIVE: To probe into the effects of electroacupuncture and salviae on expression of myocardial cellular heat-shock protein 70mRNA that acts on protecting myocardium and on dopamine level that is the mark for myocardial injury after ischemia-repeifusion as well as the interaction between electroacupuncture and salviae. DESIGN: Factorial design of two factors. SETTLNG: Department of Anesthesia of Renji Hospital Affiliated to Shanghai Second Medical Scientific University. PARTICIPANTS: The experiment was performed in Animal Experimental Room of Renji Hospital from September 2001 to December 2002, in which 24 healthy New Zealand white rabbits were employed and randomized into 4 groups, 6 rabbits in each, named ① ischemia-reperfusion group (IR group), ② electroacupuncture group (EA group), ③ salviae group (SA group), ④ electroacupuncture + salviae group (ES group). METHODS: ① IR group: ischemia-reperfusion model was prepared by clipped the anterior descend branch of cardiac coronal artery for 30 minutes and released for 2 hours. ② EA group: modeling was done as the previous method. Twenty minutes before ischemia, electroacupuncture was done on Neiguan (PC 6), Yunmen (LU 2) and Lieque (LU 7), 0.8 V in voltage, 3.0-4.0 Hz in frequency. ③ SA group: modeling was done as the previous method. Before ischemia, salviae was injected intravenously once at 1.5 mg/kg, and was injected before and after reperfusion once respectively at 1.0 mg/kg. ④ ES group: modeling was done as the previous method. Both electroacupuncture and medication were applied in the group. The content of dopamine was determined in blood before ischemia,30 minutes after ischemia and 2 hours after reperfusion successively in rabbits of every group. Reverse transcription-polymerase chain reaction (RT-PCR) was used to determine myocardial heat-shock protein 70mRNA expression in ischemic and non-ischemic areas. RESULTS: After supplemented, 24 rabbits entered result analysis. ① Expression of myocardial heat-shock protein 70mRNA: that after ischemia and reperfusion was increased significantly compared that before ischemia in every group (P ＜ 0.01). That in the rest 3 groups was all higher than IR group,and that in ES group was higher than EA and SA groups (F=4.48, P ＜0.05). CONCLUSION: ① Both electroacupuncture and salivae stimulate expression of heat-shock protein 70 mRNA after ischemia and reperfusion, enhance protein stability of heat-shock protein 70 and alleviate myocardial damage. ② Both electroacupuncture and salivae inhibit increased dopamine content in the body after ischemia and reperfusion, reduce dopamine mediated injury and protect myocardium. ③ Coordination is present between electroacupuncture and salivae.