MTDATA and the prediction of phase equilibria in oxide systems : 30 years of industrial collaboration

This paper gives an introduction to MTDATA, Phase Equilibrium Software from the National Physical Laboratory (NPL), and describes the latest advances in the development of a comprehensive database of thermodynamic parameters to underpin calculations of phase equilibria in large oxide, sulfide, and fluoride systems of industrial interest. The database, MTOX, has been developed over a period of thirty years based upon modeling work at NPL and funded by industrial partners in a project co-ordinated by Mineral Industry Research Organisation. Applications drawn from the fields of modern copper scrap smelting, high-temperature behavior of basic oxygen steelmaking slags, flash smelting of nickel, electric furnace smelting of ilmenite, and production of pure TiO2via a low-temperature molten salt route are discussed along with calculations to assess the impact of impurities on the uncertainty of fixed points used to realize the SI unit of temperature, the kelvin

Retrospective evaluation of whole exome and genome mutation calls in 746 cancer samples

Funder: NCI U24CA211006Abstract: The Cancer Genome Atlas (TCGA) and International Cancer Genome Consortium (ICGC) curated consensus somatic mutation calls using whole exome sequencing (WES) and whole genome sequencing (WGS), respectively. Here, as part of the ICGC/TCGA Pan-Cancer Analysis of Whole Genomes (PCAWG) Consortium, which aggregated whole genome sequencing data from 2,658 cancers across 38 tumour types, we compare WES and WGS side-by-side from 746 TCGA samples, finding that ~80% of mutations overlap in covered exonic regions. We estimate that low variant allele fraction (VAF < 15%) and clonal heterogeneity contribute up to 68% of private WGS mutations and 71% of private WES mutations. We observe that ~30% of private WGS mutations trace to mutations identified by a single variant caller in WES consensus efforts. WGS captures both ~50% more variation in exonic regions and un-observed mutations in loci with variable GC-content. Together, our analysis highlights technological divergences between two reproducible somatic variant detection efforts

Plant cell factories : Production of hydrophobin fusion proteins in plant cell cultures

Recombinant proteins are used as pharmaceuticals, enzymes and components of nanotechnology. The exceptional properties of fungal hydrophobins make them interesting for many of those uses. They also transfer some of their functionalities to fusion proteins enabling completely new applications. In general, plants are a potential platform for manufacturing recombinant proteins even in agricultural scale. This work explores production of hydrophobin fusion proteins in a plant cell factory: the tobacco bright yellow 2 cells (BY-2). The hydrophobin fusion technology has been mainly based on a single hydrophobin molecule the Trichoderma reesei HFBI. This work expanded the toolkit with several new molecules. When expressed in plants the hydrophobin fusion partners induced formation of protein bodies, in relation to the accumulation levels. In addition to HFBI, only HFBII and HFBIV interacted with non-ionic surfactant to selectively separate fusion proteins in surfactant based two phase separation. In Nicotiana benthamiana HFBII improved accumulation of green fluorescent protein (GFP) and Protein A in comparison to both HFBI-fused and non-fused proteins. However, HFBI, HFBII and HFBIV fusion partners all slightly reduced the yield of transferrin. Both HFBI-Protein A and transferrin-HFBIV were produced in BY-2 suspension cells with good yields. Furthermore, continuous selection resulted also in a cell line yielding 1.1 g/l GFP-HFBI. This is the first report on a plant cell culture reaching gram per litre yields. The BY-2 propagation was successfully scaled-up to 600 litre culture volume in classical stirred tank bioreactors. The aqueous two phase separation from plant cell extract was successfully scaled to 20 l volume. The fusion proteins retained functional properties from both fusion partners. The HFBI-Protein A enabled harvesting of antibodies in solution using aqueous two phase separation. The HFBIV fused transferring retained its capability to bind iron and interact with the transferrin receptor. Coating with transferrin-HFBIV resulted in uptake of the silicon nanoparticles in human cancer cells. This work builds foundation for utilization of BY-2 suspension cells in industrial manufacturing of recombinant proteins and on the other hand opens interesting new applications for bi-functional hydrophobin fusion proteins.Rekombinanttiproteiineja käytetään lääkkeinä, entsyymeinä ja nanomateriaalien komponentteina. Sieniperäisten hydrofobiinien poikkeukselliset ominaisuudet tekevät niistä kiinnostavia moniin näistä käyttökohteista. Jotkin näistä ominaisuuksista siirtyvät myös fuusioproteiineille mahdollistaen uudenlaisia sovelluksia. Kasvit ovat yleisesti lupaava tapa tuottaa rekombinanttiproteiineja jopa maatalouden suuressa mittakaavassa. Tämä tutkimus selvitti hydrofobiinifuusioproteiinien tuottamista kasvisolutehtaissa: tupakan BY-2 soluviljelmissä. Hydrofobiinifuusioteknologia on perustunut pääasiassa Trichoderma reesei-homeen HFBI molekyyliin. Tämä työ laajentaa työkalupakkia useilla molekyyleillä. Kasvisoluissa uudet hydrofobiinit muodostivat fluoresoiviin proteiineihin liitettyinä pyöreitä proteiinijyväsiä suhteessa tuottotasoon. HFBI:n lisäksi vain HFBII ja HFBIV mahdollistivat fuusioproteiinien puhdistamisen kaksifaasiuutolla. Nicotiana benthamiana-kasveissa HFBII paransi sekä fluoresoivan malliproteiinin (GFP), että proteiini A:n tuottotasoja verrattuna HFBI-fuusiopartneriin tai fuusioimattomiin proteiineihin. Kuitenkin sekä HFBI, HFBII että HFBIV huononsivat transferriinin saantoa. Sekä HFBI-Proteiini A, että transferriini-HFBIV-fuusioproteiineja tuotettiin myös BY-2 soluviljelmässä hyvin saannoin. Huolellisella valinnalla kehitettiin lisäksi solulinja, joka tuotti 1.1 g/l GFP-HFBI-proteiinia. Tämä on ensimmäinen kerta, kun kasvisoluissa on saavutettu gramma-luokan saantoja. BY-2 solujen kasvattaminen skaalattiin 600 litran tilavuuteen terästankkibioreaktoreissa. Kaksifaasiuutto kasvisolumateriaalista skaalattiin 20 litran tilavuuteen. Fuusioproteiinit säilyttivät toiminnallisia ominaisuuksia molemmilta partnereilta. HFBI-Proteiini A mahdollisti vasta-aineiden puhdistamisen liuoksista kaksifaasiuutolla. HFBIV:n liitetty transferriini taas säilytti kykynsä sitoa rautaa ja sitoutua reseptoriinsa kanssa. Nanopartikkelien pinnoittaminen transferriini-HFBIV-proteiinilla mahdollisti partikkelien kuljettamisen syöpäsoluihin. Tämä tutkimus luo pohjaa BY-2 solujen käytölle teollisessa rekombinanttiproteiinien tuotannossa ja toisaalta avaa mielenkiintoisia sovelluksia hydrofobiinifuusioproteiinien käytölle