High Density HEK293T culture for high yield, high quality, stable adenoviral vector production in Ambr® 250 stirred tank reactors

Adenovirus vectors (AdV) present high safety and immunogenicity for drug development, allowing more and more vaccines to adopt this technology platform in recent years. Due to the current COVID-19 pandemic, the global demand for AdV has experienced significant growth. Therefore, to optimize the upstream process in order to obtain high yields, good quality and stable viral vectors, it becomes critical that processes are stable and easy to scale-up, which has become a key focus of pharmaceutical companies in the field. Please click Download on the upper right corner to see the full abstract

Isolation and Characterization of a Phosphorus-Solubilizing Bacterium from Rhizosphere Soils and Its Colonization of Chinese Cabbage (Brassica campestris ssp. chinensis)

Phosphate-solubilizing bacteria (PSB) can promote the dissolution of insoluble phosphorus (P) in soil, enhancing the availability of soluble P. Thus, their application can reduce the consumption of fertilizer and aid in sustainable agricultural development. From the rhizosphere of Chinese cabbage plants grown in Yangling, we isolated a strain of PSB (YL6) with a strong ability to dissolve P and showed that this strain promoted the growth of these plants under field conditions. However, systematic research on the colonization of bacteria in the plant rhizosphere remains deficient. Thus, to further study the effects of PSB on plant growth, in this study, green fluorescent protein (GFP) was used to study the colonization of YL6 on Chinese cabbage roots. GFP expression had little effect on the ability of YL6 to grow and solubilize P. In addition, the GFP-expressing strain stably colonized the Chinese cabbage rhizosphere (the number of colonizing bacteria in the rhizosphere soil was 4.9 lg CFU/g). Using fluorescence microscopy, we observed a high abundance of YL6-GFP bacteria at the Chinese cabbage root cap and meristematic zone, as well as in the root hairs and hypocotyl epidermal cells. High quantities of GFP-expressing bacteria were recovered from Chinese cabbage plants during different planting periods for further observation, indicating that YL6-GFP had the ability to endogenously colonize the plants. This study has laid a solid and significant foundation for further research on how PSB affects the physiological processes in Chinese cabbage to promote plant growth

Correction: Root colonization and growth promotion of soybean, wheat and Chinese cabbage by Bacillus cereus YL6.

[This corrects the article DOI: 10.1371/journal.pone.0200181.]

Non-Destructive Viability Discrimination for Individual <i>Scutellaria baicalensis</i> Seeds Based on High-Throughput Phenotyping and Machine Learning

It is crucial to identify and select high-quality seeds for improving Scutellaria baicalensis yield. In this study, we present a non-destructive and accurate method for predicting Scutellaria baicalensis seed viability that used seed phenotypic data with machine-learning algorithms to distinguish between vital and dead seeds. Meanwhile, the SMOTE was used to balance the dataset and make the established viability discrimination model more efficient by avoiding problems of overfitting or under-fitting. The results showed that hyperspectral imaging (HSI) combined with detrend (DT) preprocessing and a support vector machine (SVM) model could predict Scutellaria baicalensis seed viability with a 93.3% accuracy, and increased the germination percentage of the seed lot to 99.1%, while machine vision imaging provided the highest 87.9% accuracy and 87.0% germination percentage. This strategy is suitable for large-scale Scutellaria baicalensis seed viability discrimination operations for ensuring seed quality, expanding the cultivation and production scales of Scutellaria baicalensis, and accelerating the present solving of the problem of short supply. It can help to accelerate the breeding of quality Scutellaria baicalensis varieties

A novel method to improve the removability of cone support structures in selective laser melting of 316L stainless steel

10.1016/j.jallcom.2020.157133JOURNAL OF ALLOYS AND COMPOUNDS85

Methane seepage inferred from pore water geochemistry in shallow sediments in the western slope of the Mid-Okinawa Trough

Widely distributed cold seeps are crucial sources of carbon to the seawater, whereas anaerobic oxidation of methane (AOM) and precipitation of authigenic carbonate might change the compositions of the methane-rich fluids and thus reduce the outputs of dissolved carbon to seawater. In this work, we analyze the pore water compositions of four gravity cores with high methane concentration abnormalities in pore water or in overlying seawater in the western slope of the Mid-Okinawa Trough. For the northern research area, active weak methane seepage through the seafloor is identified in the vicinity of mud volcanoes (sites C01 and C10), and strong emissions of gas and fluids may occur in the central mud volcanoes. In a submarine canyon at the south, C23 site exhibits rarely methane seepage, while high rate transportation of methane from deep sediments and associated AOM are identified at site C25 where small vertical faults and gas-bearing layers are developed at the surrounding. AOM at site C25 is indicated by (1) quasi-linearly decrease of sulfate concentrations with depths, (2) intense increases of dissolved inorganic carbon (DIC) concentrations and (3) significantly negative δ13C values (vary from −7.2 to −26.3‰) of DIC. Organoclastic degradation in the shallow sediments is not obviously recognized in the study area because of the low concentrations of NH4+, as well as negligible sulfate consumption at three of the four sites. Our data reveal that the cold systems are one of the DIC sources (the fluxes are ca. 11.2 mmol/m2/yr) to the seawater in the Okinawa Trough, which can be tracked by high depletion of 13C (δ13CDIC ranges from −7.2 to −26.3‰ VPDB)

Uterus Hyperplasia and Increased Carcinogen-Induced Tumorigenesis in Mice Carrying a Targeted Mutation of the Chk2 Phosphorylation Site in Brca1

The tumor suppressor BRCA1 contains multiple functional domains that interact with many proteins. After DNA damage, BRCA1 is phosphorylated by CHK2 at serine 988, followed by a change in its intracellular location. To study the functions of CHK2-dependent phosphorylation of BRCA1, we generated a mouse model carrying the mutation S971A (S971 in mouse Brca1 corresponds to S988 in human BRCA1) by gene targeting. Brca1(S971A/S971A) mice were born at the expected ratio without a developmental defect, unlike previously reported Brca1 mutant mice. However, Brca1(S971A/S971A) mice suffered a moderately increased risk of spontaneous tumor formation, with a majority of females developing uterus hyperplasia and ovarian abnormalities by 2 years of age. After treatment with DNA-damaging agents, Brca1(S971A/S971A) mice exhibited several abnormalities, including increased body weight, abnormal hair growth pattern, lymphoma, mammary tumors, and endometrial tumors. In addition, the onset of tumor formation became accelerated, and 80% of the mutant mice had developed tumors by 1 year of age. We demonstrated that the Brca1(S971A/S971A) cells displayed reduced ability to activate the G(2)/M cell cycle checkpoint upon γ-irradiation and to stabilize p53 following N-methyl-N′-nitro-N-nitrosoguanidine treatment. These observations suggest that Chk2 phosphorylation of S971 is involved in Brca1 function in modulating the DNA damage response and repressing tumor formation

Authigenic carbonate formation revealed by lipid biomarker inventory at hydrocarbon seeps: A case study from the Okinawa Trough

Authigenic carbonates were recovered from the northern Okinawa Trough at 540-700 m water depth. Development of microbial communities and seepage dynamics driving the precipitation of authigenic carbonates remains poorly constrained, even though the source of methane-rich fluids, formation of Fe-rich carbonates, and the potential driving forces were previously reported. Here, petrologic observations, stable carbon and oxygen isotopic compositions, mineralogy, and lipid inventories of authigenic carbonates from the northern Okinawa Trough were analyzed. The carbonate minerals were comprised predominantly of aragonite, high-magnesium calcite, and siderite. The presence of molecular fossils diagnostic for anaerobic methane oxidizing archaea (ANME) and sulfate-reducing bacteria (SRB) with pronounced C-13 depletions (as low as - 134 parts per thousand), together with highly negative delta C-13(carb) values (as low as - 55.8 parts per thousand), suggest that the carbonates precipitated from anaerobic oxidation of methane (AOM) with microbial methane as the predominant carbon source. The difference between measured and calculated delta O-18(carb) values (based on 0 parts per thousand of sea water delta O-18 vs V-SMOW) was higher than +3 parts per thousand in one of the carbonates (GGD16), reflecting an origin from gas hydrate dissociation. Most of the carbonates revealed delta O-18 offsets between measured and calculated values within + 3 parts per thousand, or a lower measured delta O-18 than calculated value, suggesting a mixture of methane-derived carbonates (low delta C-13 /high delta O-18) and detrital origin carbonates (high delta C-13/low delta O-18). Biomarker patterns varied significantly among samples. A suite of C-13-depleted biomarkers indicated the predominance of an ANME-1 assemblage for all samples, suggesting carbonates formed at low to medium methane flux. The predominance of ANME-1, abundant detrital minerals, and allochthonous biomarkers, as well as low carbonate contents indicate that most samples formed at greater depth within the sediment column. Only one carbonate sample in this study is interpreted to have precipitated at relatively shallower depth, as indicated by the occurrence of crocetane, dominance of aragonite, and higher overall carbonate content