Genetic Code Expansion System for Tight Control of Gene Expression in Bombyx mori Cell Lines

Inducible gene expression systems are important tools for studying gene function and to control protein synthesis. With the completion of the detailed map of the silkworm (Bombyx mori) genome, the study of Bombyx mori has entered the post-genome era. While the functions of many genes have been described in detail, many coding genes remain unidentified. Except for the available tetracycline induction system, there is currently a dearth of other effective induction systems for B. mori. A genetic code expansion system can be used for protein labeling and to regulate gene expression. Here, we have established a genetic code expansion system for B. mori based on the well-researched tRNAPyl/PylRS pair from Methanosarcina mazei. We used H-Lys(Boc)-OH, which is a lysine derivative to efficiently and tightly control the expression of the reporter gene DsRed[TAG]EGFP (D[TAG]G), which encoded a H-Lys(Boc)-OH-bearing protein fused with DsRed and EGFP (here regarded as D[Boc]G) in B. mori cell lines BmE and BmNs. In D[TAG]G, the amber stop codon is recognized as the orthogonal tRNAPyl. Successful application of genetic code expansion system in silkworm cell lines will support the research into the function of silkworm genes and paves the way for the identification of new genes and protein markers in silkworm.

Analysis of the mathematical modelling of a static expansion system

Static expansion systems are used to generate pressures in medium and high vacuum and are used in the calibration of absolute pressure meters in these pressure ranges. In the present study, the suitability of different models to represent the final pressures in a static expansion system with two tanks is analysed. It is concluded that the use of the ideal gas model is adequate in most simulated conditions, while the assumption that the residual pressure is zero before expansion presents problems under certain conditions. An uncertainty analysis of the process is carried out, which leads to evidence of the high importance of uncertainty in a first expansion over subsequent expansion processes. Finally, an analysis of the expansion system based on uncertainty is carried out to estimate the effect of the metrological characteristics of the measurements of the input quantities. Said design process can make it possible to determine a set of restrictions on the uncertainties of the input quantities.

Influence of Starch Content on the Thermal and Viscoelastic Properties of Syndiotactic Polypropylene/Starch Composites

In this study, syndiotactic Polypropylene/Starch (sPP/starch) composites were prepared using a solution mixing technique. The thermal characterization was performed using Differential Scanning Calorimetry (DSC), and the melting point was measured for all polymer composites. The thermal degradation temperature was measured using thermal gravimetric analysis. The viscoelastic measurements were performed using the Atomic Rheometric Expansion System (ARES). Both melting point and thermal degradation temperatures were found to decrease with increasing starch content. Moreover, the elastic modulus was found to decrease when the starch content increased.

Helios is a marker, not a driver, of human Treg stability

Regulatory T cell (Treg) therapy holds promise as a potentially curative approach to establish immune tolerance in transplantation and autoimmune disease. An outstanding question is whether therapeutic Tregs have the potential to transdifferentiate into effector T cells and thus exacerbate rather than suppress immune responses. In mice, the transcription factor Helios is thought to promote Tregs lineage stability in a range of inflammatory contexts. In humans, the role of Helios in Tregs is less clear, in part due to the inability to enrich and study subsets of Helios-positive versus Helios-negative Tregs. Using an in vitro expansion system, we found that loss of high Helios expression and emergence of an intermediate Helios (Heliosmid)-expressing population correlated with Treg destabilization. We then used CRISPR/Cas9 to genetically ablate Helios expression in human naive or memory Tregs and found that Helios-knockout and unedited Tregs were equivalent in their suppressive function and stability in inflammation. Thus, high Helios expression is a marker, but not a driver, of human Treg stability in vitro. These data highlight the importance of monitoring Helios expression in therapeutic Treg manufacturing and provide new insight into the biological function of this transcription factor in human T cells.

Characterization of spinning rotor gauge-3 using orific flow system and static expansion system

This article elucidates the calibration of newly procured spinning rotor gauge (SRG 3) from MKS Instruments, USA using primary vacuum standards: Orifice flow system (OFS) and Static expansion system (SES) established at National Physical Laboratory, India (NPLI) in the range of 10^-4 Pa to 1 Pa and further compared with manufacturer reported value which was calibrated by transfer standard of National Institute of Standards & Technology (NIST). The key parameters to calculate the pressure measured by SRG is the accommodation coefficients. The accommodation coefficients for N₂ gas obtained using OFS, SES, and calibration report form NIST USA (SRG2) are 0.957, 0.961, and 0.954 respectively.