Тransplastomic tobacco plants producing the hydrophilic domain of the sheep pox virus coat protein L1R

Sheep pox has a wide geographical range of distribution and poses a threat to sheep breeding worldwide, as the disease is highly contagious and is accompanied by large economic losses. Vaccines based on live attenuated virus strains are currently being used for prevention of this disease. Such vaccines are effective, but potentially dangerous because of the possible virus reversion to a pathogenic state. The development of safe recombinant subunit vaccines against sheep pox is very relevant. The high ploidy level of the plant chloroplasts makes it possible to obtain large quantities of foreign proteins. The purpose of this study was to create transplastomic Nicotiana tabacum plants producing one of the candidate vaccine proteins of sheep pox virus L1R. A vector containing a deletion variant of the SPPV_56 gene, which encodes the N-terminal hydrophilic part of the viral coat protein L1R, was constructed to transform tobacco plastids. It provides integration of the transgene into the trnG/trnfM region of the chloroplast tobacco genome by homologous recombination. Spectinomycin-resistant tobacco lines were obtained by biolistic gun-mediated genetic transformation. PCR analysis in the presence of gene-specific primers confirmed integration of the transgene into the plant genome. Subsequent Northern and Western blot analysis showed the gene expression at the transcriptional and translational levels. The recombinant protein yields reached up to 0.9 % of total soluble protein. The transplastomic plants displayed a growth retardation and pale green leaf color compared to the wild type, but they developed normally and produced seeds. Southern blot analysis showed heteroplasmy of the plastids in the obtained plants due to recombination events between native and introduced regulatory plastid DNA elements. The recombinant protein from plant tissue was purified using metal affinity chromatography. Future research will be focused on determining the potential of the chloroplast-produced protein to induce neutralizing antibodies against SPPV strains

Growth of 3C-SiC Films on Si (111) and Sapphire (0001) Substrates by MOCVD

Thick silicon carbide films were grown on sapphire (0001) and silicon (111) substrates using metal organic chemical vapor deposition (MOCVD). Diethylmethylsilane (DEMS) has been used as a single precursor, which contain Si and C atoms in the same molecule, without any carrier or bubbler gas. Atomic structure, surface composition and morphology have been investigated by XRD, AES, SEM and AFM analysis. SiC films of 5-7 micron thickness were grown at a rate of ~ 40 nm/min on sapphire (0001) and Si (111) substrates. The films grown at low temperature (850 ºC and 900 ºC) on both substrates show crystalline 3C-SiC in the (111) orientation. XRD results show that the orientation of the crystal structure does not depend of the substrate orientation AFM pictures of SiC films grown on sapphire (0001) exhibit more crystalline order as compared to films grown on the Si (111) substrates. AES of the grown films shows that in both cases the Si peak intensity is greater than that of carbon. This work shows promise for the development of alternative processes for developing low cost, large area substrates for application to IIInitrides LED and UV photodetector fabrication and also for gas detector application

Constructing the constitutively active ribosomal protein S6 kinase 2 from <i>Arabidopsis thaliana</i> (AtRPS6K2) and testing its activity <i>in vitro</i>

Ribosomal protein S6 (RPS6) is the only phosphorylatable protein of the eukaryotic 40S ribosomal subunit. Ribosomes with phosphorylated RPS6 can selectively translate 5’TOP-(5’-terminal oligopyrimidine)-containing mRNAs that encode most proteins of the translation apparatus. The study of translational control of 5’TOP-mRNAs, which are preferentially translated when RPS6 is phosphorylated and cease to be translated when RPS6 is de-phosphorylated, is particularly important. In Arabidopsis thaliana, AtRPS6 is phosphorylated by kinase AtRPS6K2, which should in turn be phosphorylated by upper level kinases (AtPDK1 – at serine (S) 296, AtTOR – at threonine (T) 455 and S437) for full activation. We have cloned AtRPS6K2 cDNA gene and carried out in vitro mutagenesis replacing codons encoding S296, S437 and T455 by triplets of phosphomimetic glutamic acid (E). After the expression of both natural and mutated cDNAs in Escherichia coli cells, two recombinant proteins were isolated: native AtRPS6K2 and presumably constitutively active AtRPS6K2(S296E, S437E, T455E). The activity of these variants was tested in vitro. Both kinases could phosphorylate wheat (Triticum aestivum L.) TaRPS6 as part of 40S ribosomal subunits isolated from wheat embryos, though the non-mutated variant had less activity than phosphomimetic one. The ability of recombinant non-mutated kinase to phosphorylate TaRPS6 can be explained by its phosphorylation by bacterial kinases during the expression and isolation steps. The phosphomimetically mutated AtRPS6K2(S296E, S437E, T455E) can serve as a tool to investigate preferential translation of 5’TOP-mRNAs in wheat germ cell-free system, in which most of 40S ribosomal subunits have phosphorylated TaRPS6. Besides, such an approach has a biotechnological application in producing genetically modified plants with increased biomass and productivity through stimulation of cell growth and division

Ancient genomic time transect from the Central Asian Steppe unravels the history of the Scythians

The Scythians were a multitude of horse-warrior nomad cultures dwelling in the Eurasian steppe during the first millennium BCE. Because of the lack of first-hand written records, little is known about the origins and relations among the different cultures. To address these questions, we produced genome-wide data for 111 ancient individuals retrieved from 39 archaeological sites from the first millennia BCE and CE across the Central Asian Steppe. We uncovered major admixture events in the Late Bronze Age forming the genetic substratum for two main Iron Age gene-pools emerging around the Altai and the Urals respectively. Their demise was mirrored by new genetic turnovers, linked to the spread of the eastern nomad empires in the first centuries CE. Compared to the high genetic heterogeneity of the past, the homogenization of the present-day Kazakhs gene pool is notable, likely a result of 400 years of strict exogamous social rules.Introduction Results - The IA transition in the Kazakh Steppe - Admixture modeling of IA steppe populations - Post-IA genetic turnovers in the Kazakh Steppe - Dating ancient admixture - Present-day Kazakhs Discussion Material and methods - Radiocarbon dating - DNA extraction, library preparations, and sequencing - Modern DNA genotyping and quality controls - Ancient DNA data processing -- Raw data -- Authentication and contamination estimate -- Genotyping -- Sex determination -- Genetic relatedness estimation - Uniparental haplogroup assignment - Population structure analyses - Individual labeling and population grouping criteria - F-statistics and ancestry modeling - Admixture dating - CHROMOPAINTER and fineSTRUCTURE analyse

Stone Age Yersinia pestis genomes shed light on the early evolution, diversity, and ecology of plague

The bacterial pathogenYersinia pestisgave rise to devastating outbreaks throughouthuman history, and ancient DNA evidence has shown it afflicted human populations asfar back as the Neolithic.Y. pestisgenomes recovered from the Eurasian Late Neolithic/Early Bronze Age (LNBA) period have uncovered key evolutionary steps that led to itsemergence from aYersinia pseudotuberculosis-like progenitor; however, the number ofreconstructed LNBA genomes are too few to explore its diversity during this criticalperiod of development. Here, we present 17Y. pestisgenomes dating to 5,000 to 2,500y BP from a wide geographic expanse across Eurasia. This increased dataset enabled usto explore correlations between temporal, geographical, and genetic distance. Ourresults suggest a nonflea-adapted and potentially extinct single lineage that persistedover millennia without significant parallel diversification, accompanied by rapid dis-persal across continents throughout this period, a trend not observed in other pathogensfor which ancient genomes are available. A stepwise pattern of gene loss provides fur-ther clues on its early evolution and potential adaptation. We also discover the presenceof theflea-adapted form ofY. pestisin Bronze Age Iberia, previously only identified inin the Caucasus and the Volga regions, suggesting a much wider geographic spread ofthis form ofY. pestis. Together, these data reveal the dynamic nature of plague’s forma-tive years in terms of its early evolution and ecology

Stone Age Yersinia pestis genomes shed light on the early evolution, diversity, and ecology of plague

[Significance] The bacterium Yersinia pestis has caused numerous historically documented outbreaks of plague and research using ancient DNA could demonstrate that it already affected human populations during the Neolithic. However, the pathogen’s genetic diversity, geographic spread, and transmission dynamics during this early period of Y. pestis evolution are largely unexplored. Here, we describe a set of ancient plague genomes up to 5,000 y old from across Eurasia. Our data demonstrate that two genetically distinct forms of Y. pestis evolved in parallel and were both distributed across vast geographic distances, potentially occupying different ecological niches. Interpreted within the archeological context, our results suggest that the spread of plague during this period was linked to increased human mobility and intensification of animal husbandry.The bacterial pathogen Yersinia pestis gave rise to devastating outbreaks throughout human history, and ancient DNA evidence has shown it afflicted human populations as far back as the Neolithic. Y. pestis genomes recovered from the Eurasian Late Neolithic/Early Bronze Age (LNBA) period have uncovered key evolutionary steps that led to its emergence from a Yersinia pseudotuberculosis-like progenitor; however, the number of reconstructed LNBA genomes are too few to explore its diversity during this critical period of development. Here, we present 17 Y. pestis genomes dating to 5,000 to 2,500 y BP from a wide geographic expanse across Eurasia. This increased dataset enabled us to explore correlations between temporal, geographical, and genetic distance. Our results suggest a nonflea-adapted and potentially extinct single lineage that persisted over millennia without significant parallel diversification, accompanied by rapid dispersal across continents throughout this period, a trend not observed in other pathogens for which ancient genomes are available. A stepwise pattern of gene loss provides further clues on its early evolution and potential adaptation. We also discover the presence of the flea-adapted form of Y. pestis in Bronze Age Iberia, previously only identified in in the Caucasus and the Volga regions, suggesting a much wider geographic spread of this form of Y. pestis. Together, these data reveal the dynamic nature of plague’s formative years in terms of its early evolution and ecology.This study was funded by the Max Planck Society, Max Planck Harvard Research Center for the Archaeoscience of the Ancient Mediterranean and the European Research Council under the European Union’s Horizon 2020 research and innovation program under Grant Agreement 771234 – PALEoRIDER (to W.H.), 856453 – HistoGenes (to J.K.), and 834616 – ARCHCAUCASUS (to S.H.). The Heidelberg Academy of Science financed the genetic and archeological research on human individuals from the Augsburg region within the project WIN Kolleg: “Times of Upheaval: Changes of Society and Landscape at the Beginning of the Bronze Age. M.E. was supported by the award “Praemium Academiae” of the Czech Academy of Sciences. M.D. was supported by the project RVO 67985912 of the Institute of Archaeology of the Czech Academy of Sciences, Prague. I.O. was supported by the Ramón y Cajal grant from Ministerio de Ciencia e Innovación, Spanish Government (RYC2019-027909-I). A. H€ubner was supported by the Deutsche Forschungsgemeinschaft under Germany’s Excellence Strategy (EXC 2051 – Project-ID 390713860). J.F.-E. and J.A.M.-A. were supported by the Diputación Foral de Alava, IT 1223-19, Gobierno Vasco. A. Buzhilova was supported by the Center of Information Technologies and Systems (CITIS), Moscow, Russia 121041500329-0. L. M., L.B.D., and E. Khussainova were supported by the Grant AP08856654, Ministry of Education and Science of the Republic of Kazakhstan. A. Beisenov was supported by the Grant AP08857177, Ministry of Education and Science of the Republic of Kazakhstan.Peer reviewe

137 ancient human genomes from across the Eurasian steppes

Descriptive and Comparative Linguistic

Ten millennia of hepatitis B virus evolution

Hepatitis B virus (HBV) has been infecting humans for millennia and remains a global health problem, but its past diversity and dispersal routes are largely unknown. We generated HBV genomic data from 137 Eurasians and Native Americans dated between similar to 10,500 and similar to 400 years ago. We date the most recent common ancestor of all HBV lineages to between similar to 20,000 and 12,000 years ago, with the virus present in European and South American hunter-gatherers during the early Holocene. After the European Neolithic transition, Mesolithic HBV strains were replaced by a lineage likely disseminated by early farmers that prevailed throughout western Eurasia for similar to 4000 years, declining around the end of the 2nd millennium BCE. The only remnant of this prehistoric HBV diversity is the rare genotype G, which appears to have reemerged during the HIV pandemic.Molecular Technology and Informatics for Personalised Medicine and Healt

Ten millennia of hepatitis B virus evolution

Hepatitis B virus (HBV) has been infecting humans for millennia and remains a global health problem, but its past diversity and dispersal routes are largely unknown. We generated HBV genomic data from 137 Eurasians and Native Americans dated between ~10,500 and ~400 years ago. We date the most recent common ancestor of all HBV lineages to between ~20,000 and 12,000 years ago, with the virus present in European and South American hunter-gatherers during the early Holocene. After the European Neolithic transition, Mesolithic HBV strains were replaced by a lineage likely disseminated by early farmers that prevailed throughout western Eurasia for ~4000 years, declining around the end of the 2nd millennium BCE. The only remnant of this prehistoric HBV diversity is the rare genotype G, which appears to have reemerged during the HIV pandemic