Optical and magneto-optical properties of single crystals of RFe2 (R=Gd, Tb, Ho, and Lu) and GdCo2 intermetallic compounds

We have studied the diagonal and off-diagonal optical conductivity of a RFe2(R=Gd, Tb, Ho, Lu) and GdCo2 single crystal grown by the flux method. Using spectroscopic ellipsometry we have measured the dielectric function from 1.5 to 5.5 eV. The magneto-optical parameters (Kerr rotation and ellipticity) were obtained using a magneto-optical polar Kerr spectrometer at temperatures between 7 and 295 K and applied magnetic fields between 0.5 to 1.6 T. The apparatus and calibration method are described in detail. Using magneto-optical data and optical constants we derive the experimental value of the off-diagonal conductivity components.;Theoretical calculations of optical conductivities and magneto-optical parameters were performed using the tight binding-linear muffin tin orbitals method within the local spin density approximation. We applied this TB-LMTO method to LuFe2. The obtained theoretical results agree well with the experimental data. The oxidation effects on the diagonal part of the optical conductivity were considered using a three phase model. The oxidation effects on the magneto-optical parameters were also considered by treating the oxide layer as a nonmagnetic thin transparent layer. These corrections change not only the magnitude but also the shape of the optical conductivity and the magneto-optical parameters

The electron temperature of the inner halo of the Planetary Nebula NGC 6543

We investigate the electron temperature of the inner halo and nebular core regions of NGC 6543, using archival Hubble Space Telescope (HST) Wide Field Planetary Camera 2 (WFPC2) images taken through narrow band [O III] filters. Balick et al. (2001) showed that the inner halo consists of a number of spherical shells. We find the temperature of this inner halo to be much higher (~15000 K) than that of the bright core nebula (~8500 K). Photo-ionization models indicate that hardening of the UV radiation from the central star cannot be the main source of the higher temperature in the halo region. Using a radiation hydrodynamic simulation, we show that mass loss and velocity variations in the AGB wind can explain the observed shells, as well as the higher electron temperature.Comment: 9 pages, 6 figures, to be published in A&

Genomic understanding of clonal variation in recombinant CHO cells

In the current manufacturing platform for large-scale production of therapeutic proteins, recombinant Chinese hamster ovary (rCHO) cell line generation is probably the most time-consuming step. The high degree of phenotypic heterogeneity in the absolute transgene expression level and variable responses to culture conditions among the selected clones, which has been referred to as “clonal variation”, makes the cell line generation process laborious and time-consuming. Upon transfection, a transgene is randomly integrated into the chromosomes of the host cells, and the integration site of the transgene is believed to be responsible for this clonal variation. However, the lack of understanding the molecular basis involved in clonal variation has hindered rapid cell line generation with a predicted culture performance. With the availability of CHO genome sequences and targeted genome sequencing/editing technologies, we investigated “clonal variation” with an emphasis on the transgene integration sites and configuration of the integrated transgenes. rCHO cell clones expressing GFP, which were constructed by random integration of the GFP expression vector into CHOK1 host cells, revealed a remarkable variability in GFP expression at 37°C. Furthermore, they showed different responses to culture temperature shifts from 37°C to 33°C confirming the existence of clonal variation. Application of targeted sequencing by the proximity ligation to the transgenes enabled the mapping of the integrated transgenes, and thereby the genomic integration sites of the transgene in the representative rCHO cell clones showing different responses to hypothermia were identified. To determine whether the different responses of the rCHO clones to hypothermia were due to the different integration sites of the transgenes, rCHO cell clones expressing GFP were also constructed by CRISPR/Cas9-mediated targeted integration of an intact transgene into CHOK1 host cells at the integration sites that were identified in the rCHO cells exhibiting different responses to hypothermia. Surprisingly, the rCHO clones constructed with targeted integration, regardless of the integration site of the gene, had similar expression patterns in terms of the absolute expression level and responses to hypothermia. They all exhibited enhanced GFP expression with hypothermia. Evidence of several rearrangements in the integrated transgene was detected in the rCHO clones constructed with random integration, which may interrupt the normal function of regulatory elements, particularly promoters. Promoter replacement and dissection results support the crucial role of promoter elements in the differential transgene expression patterns at the identical genomic site. Taken together, we demonstrate the complex nature of “clonal variation” in rCHO cells which encompasses the concept of vector elements and their rearrangement upon random integration besides just the genomic integration sites. Further characterization of the interactions between the integration sites and vector regulatory elements together with controlled integration of transgenes could lead to the tailored control of recombinant gene expression in rCHO cells while minimizing clonal variation