A versatile reverse genetics platform for SARS-CoV-2 and other positive-strand RNA viruses

The current COVID-19 pandemic is caused by the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2). We demonstrate that despite the large size of the viral RNA genome (~30 kb), infectious full-length cDNA is readily assembled in vitro by a circular polymerase extension reaction (CPER) methodology without the need for technically demanding intermediate steps. Overlapping cDNA fragments are generated from viral RNA and assembled together with a linker fragment containing CMV promoter into a circular full-length viral cDNA in a single reaction. Transfection of the circular cDNA into mammalian cells results in the recovery of infectious SARS-CoV-2 virus that exhibits properties comparable to the parental virus in vitro and in vivo. CPER is also used to generate insect-specific Casuarina virus with ~20 kb genome and the human pathogens Ross River virus (Alphavirus) and Norovirus (Calicivirus), with the latter from a clinical sample. Additionally, reporter and mutant viruses are generated and employed to study virus replication and virus-receptor interactions

Textural history of recent basaltic-andesites and plutonic inclusions from Merapi volcano

Mt. Merapi in Central Java is one of the most active stratovolcanoes on Earth and is underlain by a multistage plumbing system. Crystal size distribution analyses (CSD) were carried out on recent Merapi basaltic-andesites and co-eruptive magmatic and plutonic inclusions to characterise the crystallisation processes that operate during storage and ascent and to obtain information on respective time scales. The basaltic-andesites exhibit log-linear, kinked-upwards CSD curves for plagioclase and clinopyroxene that can be separated into two main textural populations. Large plagioclase phenocrysts (≥1.6 mm) make up one population, but correspond to crystals with variable geochemical composition and reflect a period of crystal growth at deep to mid-crustal levels. This population was subsequently influenced by crystal accumulation and the onset of crustal assimilation, including the incorporation of high-Ca skarn-derived xenocrysts. Textural re-equilibration is required for these crystals to form a single population in CSD. A second episode of crystal growth at shallower levels is represented by chemically homogenous plagioclase crystals <1.6 mm in size. Crustal assimilation is indicated by, for example, oxygen isotopes and based on the CSD data, crystallisation combined with contamination is likely semi-continuous in these upper crustal storage chambers. The CSD data observed in the basaltic-andesite samples are remarkably consistent and require a large-volume steady state magmatic system beneath Merapi in which late textural equilibration plays a significant role. Plagioclase CSDs of co-eruptive magmatic and plutonic inclusions may contain a third crystal population (<1 mm) not found in the lavas. This third population has probably formed from enhanced degassing of portions of basaltic-andesite magma at shallow crustal levels which resulted in increased crystallinity and basaltic-andesite mush inclusions. A suite of coarse plutonic inclusions is also present that reflects crystallisation and accumulation of crystals in the deep Merapi plumbing system, as deduced from CSD patterns and mineral assemblages