Expression and purification of soluble single-chain Fv against human fibroblast growth factor receptor 3 fused with Sumo tag in Escherichia coli

Background: Overexpression or mutated activation of Fibroblast growth factor receptor 3 (FGFR3) is involved in the pathogenesis of many tumors. More and more studies focus on the potential usage of therapeutic antibodies against FGFR3. Results: In this study, a novel single-chain Fv (ScFv) against FGFR3 was prepared and characterized. To achieve the soluble expression, ScFv was fused with Sumo (Small ubiquitin-related modifier) by polymerase chain reaction (PCR), and cloned into pET-20b. The recombinant bacteria were induced by 0.5 mM Isopropyl-\u3b2-D-thiogalactopyranoside (IPTG) for 16 h at 20\ub0C, and the supernatant liquid of Sumo-ScFv was harvested and purified by Ni-NTA chromatography. After being cleaved by the Sumo protease, the recombinant ScFv was released from the fusion protein, and further purified by Ni-NTA chromatography. The purity of ScFv was shown to be higher than 95% and their yield reached 4 mg per liter of bacterial culture. In vitro data showed that ScFv can significantly attenuate FGF9-induced phosphorylation of FGFR3. Conclusion: We provide a novel method to produce soluble expression and bioactive functions of ScFv in Escherichia coli

Robust estimation of bacterial cell count from optical density

Optical density (OD) is widely used to estimate the density of cells in liquid culture, but cannot be compared between instruments without a standardized calibration protocol and is challenging to relate to actual cell count. We address this with an interlaboratory study comparing three simple, low-cost, and highly accessible OD calibration protocols across 244 laboratories, applied to eight strains of constitutive GFP-expressing E. coli. Based on our results, we recommend calibrating OD to estimated cell count using serial dilution of silica microspheres, which produces highly precise calibration (95.5% of residuals <1.2-fold), is easily assessed for quality control, also assesses instrument effective linear range, and can be combined with fluorescence calibration to obtain units of Molecules of Equivalent Fluorescein (MEFL) per cell, allowing direct comparison and data fusion with flow cytometry measurements: in our study, fluorescence per cell measurements showed only a 1.07-fold mean difference between plate reader and flow cytometry data

Thymosin beta 10 Prompted the VEGF-C Expression in Lung Cancer Cell

Background and objective Our previous study found that thymosin β10 overexpressed in lung cancer and positively correlated with differentiation, lymph node metastasis and stage of lung cancer. In this reasearch we aim to study the effects and mechanism of exogenous human recombinant Tβ10 on the expression of VEGF-C on non-small cell lung cancer. Methods After SPC, A549 and LK2 cells were treated with 100 ng/mL recombinant human Tβ10, the mRNA level of VEGF-C were detected by RT-PCR. The mean while the protein expression of VEGF-C, P-AKT and AKT were determined by Western blot assay. Results Exogenous recombinant human Tβ10 were significantly promote the expression levels of VEGF-C mRNA and protein while promoting the phosphorylation of AKT. Exogenous Tβ10 can promote the expression of VEGF-C mRNA and protein in lung cancer cell lines A549 and LK2 (P<0.05), and this effect can be inhibited by use AKT inhibitor LY294002 (P<0.05). Conclusion Tβ10 human recombinant proteins can promote the expression of VEGF-C by activating AKT phosphorylation in lung cancer cell lines

Application of UAV Photogrammetric System for Monitoring Ancient Tree Communities in Beijing

Ancient tree community surveys have great scientific value to the study of biological resources, plant distribution, environmental change, genetic characteristics of species, and historical and cultural heritage. The largest ancient pear tree communities in China, which are rare, are located in the Daxing District of Beijing. However, the environmental conditions are tough, and the distribution is relatively dispersed. Therefore, a low-cost, high-efficiency, and high-precision measuring system is urgently needed to complete the survey of ancient tree communities. By unmanned aerial vehicle (UAV) photogrammetric program research, ancient tree information extraction method research, and ancient tree diameter at breast height (DBH) and age prediction model research, the proposed method can realize the measurement of tree height, crown width, and prediction of DBH and tree age with low cost, high efficiency, and high precision. Through experiments and analysis, the root mean square error (RMSE) of the tree height measurement was 0.1814 m, the RMSE of the crown width measurement was 0.3292 m, the RMSE of the DBH prediction was 3.0039 cm, and the RMSE of the tree age prediction was 4.3753 years, which could meet the needs of ancient tree survey of the Daxing District Gardening and Greening Bureau. Therefore, a UAV photogrammetric measurement system proved to be capable when applied in the survey of ancient tree communities and even in partial forest inventories

Chemical Structures and Pharmacological Properties of Typical Bioflavonoids in Polygonati Rhizoma (PGR)

Most medicines are coming with toxic and detrimental side effects. In addition, microbials are resisting the medicine. Therefore, alternative drugs with low toxic and side effects and low microbial resistance are needed. Plants offer good potential candidates due to a broad range of chemicals they contain. These chemicals have been studied, and research is still going on to probe chemical properties of plant chemicals. In China, traditional Chinese medicine is practised, whereby plant extracts are obtained, and then sold in packages for reasons like memory enhancement, cancer treatment, boosting immune system, and so on. Among the herbs cultivated in China is Polygonati rhizoma (PGR). This plant contains various bioflavonoids such as diosgenin, kaempferol, catechin, daidzein, and 3′-methoxydaidzein. In this review, we discussed the pharmacological effects of these chemicals, including luteolin antimicrobial activity in a manner that it circumvents antibiotic resistance; rutin antivenom property; kaempferol as an agent that mitigates neuropathic pain; genistein anticancer property; isorhamnetin’s ability to alleviate chronic obstructive pulmonary diseases (COPD); proanthocyanidins’ ability to deal with diabetic neuropathy and analgesic property of catechin