De novo assembly of RNA-seq data enables researchers to study transcriptomes without the need for a genome sequence; this approach can be usefully applied, for instance, in research on 'non-model ...organisms' of ecological and evolutionary importance, cancer samples or the microbiome. In this protocol we describe the use of the Trinity platform for de novo transcriptome assembly from RNA-seq data in non-model organisms. We also present Trinity-supported companion utilities for downstream applications, including RSEM for transcript abundance estimation, R/Bioconductor packages for identifying differentially expressed transcripts across samples and approaches to identify protein-coding genes. In the procedure, we provide a workflow for genome-independent transcriptome analysis leveraging the Trinity platform. The software, documentation and demonstrations are freely available from http://trinityrnaseq.sourceforge.net. The run time of this protocol is highly dependent on the size and complexity of data to be analyzed. The example data set analyzed in the procedure detailed herein can be processed in less than 5 h.
Abstract
The Human Proteoform Atlas (HPfA) is a web-based repository of experimentally verified human proteoforms on-line at http://human-proteoform-atlas.org and is a direct descendant of the ...Consortium of Top-Down Proteomics’ (CTDP) Proteoform Atlas. Proteoforms are the specific forms of protein molecules expressed by our cells and include the unique combination of post-translational modifications (PTMs), alternative splicing and other sources of variation deriving from a specific gene. The HPfA uses a FAIR system to assign persistent identifiers to proteoforms which allows for redundancy calling and tracking from prior and future studies in the growing community of proteoform biology and measurement. The HPfA is organized around open ontologies and enables flexible classification of proteoforms. To achieve this, a public registry of experimentally verified proteoforms was also created. Submission of new proteoforms can be processed through email vianrtdphelp@northwestern.edu, and future iterations of these proteoform atlases will help to organize and assign function to proteoforms, their PTMs and their complexes in the years ahead.
A full description of the human proteome relies on the challenging task of detecting mature and changing forms of protein molecules in the body. Large-scale proteome analysis has routinely involved ...digesting intact proteins followed by inferred protein identification using mass spectrometry. This 'bottom-up' process affords a high number of identifications (not always unique to a single gene). However, complications arise from incomplete or ambiguous characterization of alternative splice forms, diverse modifications (for example, acetylation and methylation) and endogenous protein cleavages, especially when combinations of these create complex patterns of intact protein isoforms and species. 'Top-down' interrogation of whole proteins can overcome these problems for individual proteins, but has not been achieved on a proteome scale owing to the lack of intact protein fractionation methods that are well integrated with tandem mass spectrometry. Here we show, using a new four-dimensional separation system, identification of 1,043 gene products from human cells that are dispersed into more than 3,000 protein species created by post-translational modification (PTM), RNA splicing and proteolysis. The overall system produced greater than 20-fold increases in both separation power and proteome coverage, enabling the identification of proteins up to 105 kDa and those with up to 11 transmembrane helices. Many previously undetected isoforms of endogenous human proteins were mapped, including changes in multiply modified species in response to accelerated cellular ageing (senescence) induced by DNA damage. Integrated with the latest version of the Swiss-Prot database, the data provide precise correlations to individual genes and proof-of-concept for large-scale interrogation of whole protein molecules. The technology promises to improve the link between proteomics data and complex phenotypes in basic biology and disease research.
Full text
Available for:
DOBA, IJS, IZUM, KILJ, NUK, PILJ, PNG, SAZU, SIK, UILJ, UKNU, UL, UM, UPUK
A proteoform is a defined form of a protein derived from a given gene with a specific amino acid sequence and localized post‐translational modifications. In top‐down proteomic analyses, proteoforms ...are identified and quantified through mass spectrometric analysis of intact proteins. Recent technological developments have enabled comprehensive proteoform analyses in complex samples, and an increasing number of laboratories are adopting top‐down proteomic workflows. In this review, some recent advances are outlined and current challenges and future directions for the field are discussed.
Full text
Available for:
BFBNIB, FZAB, GIS, IJS, KILJ, NLZOH, NUK, OILJ, SAZU, SBCE, SBMB, UL, UM, UPUK
After decades of declining cropland area, the United States (US) experienced a reversal in land use/land cover change in recent years, with substantial grassland conversion to cropland in the US ...Midwest. Although previous studies estimated soil carbon (C) loss due to cropland expansion, other important environmental indicators, such as soil erosion and nutrient loss, remain largely unquantified. Here, we simulated environmental impacts from the conversion of grassland to corn and soybeans for 12 US Midwestern states using the EPIC (Environmental Policy Integrated Climate) model. Between 2008 and 2016, over 2 Mha of grassland were converted to crop production in these states, with much less cropland concomitantly abandoned or retired from production. The net change in grassland-cropland conversion increased annual soil erosion by 7.9%, nitrogen (N) loss by 3.7%, and soil organic carbon loss by 5.6% relative to that of existing cropland, despite an associated increase in cropland area of only 2.5%. Notably, the above estimates represent the scenario of converting unmanaged grassland to tilled corn and soybeans, and impacts varied depending upon crop type and tillage regime. Corn and soybeans are dominant biofuel feedstocks, yet the grassland conversion and subsequent environmental impacts simulated in this study are likely not attributable solely to biofuel-driven land use change since other factors also contribute to corn and soybean prices and land use decisions. Nevertheless, our results suggest grassland conversion in the Upper Midwest has resulted in substantial degradation of soil quality, with implications for air and water quality as well. Additional conservation measures are likely necessary to counterbalance the impacts, particularly in areas with high rates of grassland conversion (e.g., the Dakotas, southern Iowa).
The Conservation Reserve Program (CRP) is the largest agricultural land-retirement program in the United States, providing many environmental benefits, including wildlife habitat and improved air, ...water, and soil quality. Since 2007, however, CRP area has declined by over 25% nationally with much of this land returning to agriculture. Despite this trend, it is unclear what types of CRP land are being converted, to what crops, and where. All of these specific factors greatly affect environmental impacts. To answer these questions, we quantified shifts in expiring CRP parcels to five major crop-types (corn, soy, winter and spring wheat, and sorghum) in a 12-state, Midwestern region of the United States using a US Department of Agriculture (USDA), field-level CRP database and USDA's Cropland Data Layer. For the years 2010 through 2013, we estimate almost 30%, or more than 530 000 ha, of expiring CRP land returned to the production of these five crops in our study area, with soy and corn accounting for the vast majority of these shifts. Grasslands were the largest type of CRP land converted (360 000 ha), followed by specifically designated wildlife habitat (76 000 ha), and wetland areas (53 000 ha). These wetland areas were not just wetlands themselves, but also a mix of land covers enhancing or protecting wetland ecosystem services (e.g., wetland buffers). Areas in the Dakotas, Nebraska, and southern Iowa were hotspots of change, with the highest areas of CRP land moving back to agriculture. By contrast, we estimate only a small amount (∼3%) of the expiring land shifted into similar, non-CRP land-retirement or easement programs. Reconciling needs for food, feed, fuel, and healthy ecosystems is an immense challenge for farmers, conservationists, and state and federal agencies. Reduced enrollment and the turnover of CRP land from conservation to agriculture raises questions about sustaining ecosystem services in this region.
The Touch And Go Camera System (TAGCAMS) is a three-camera-head instrument onboard NASA’s OSIRIS-REx asteroid sample return mission spacecraft. The purpose of TAGCAMS is to facilitate navigation to ...the target asteroid, (101955) Bennu; confirm acquisition of the asteroid sample; document asteroid sample stowage; and provide supplementary imaging for OSIRIS-REx science investigations.
During the almost two-year OSIRIS-REx outbound cruise phase we pursued nine TAGCAMS imaging campaigns to check, calibrate and characterize the camera system’s performance before asteroid arrival and proximity operations began in late 2018. The TAGCAMS in-flight calibration dataset provides the relevant information to enable the three cameras to complete their primary observation goals during asteroid operations. The key performance parameters that we investigated in flight included: linearity, responsivity (both point source and extended body), dark current, hot pixels, pointing, image geometry transformation, image quality and stray light. Analyses of the in-flight performance either confirmed the continued applicability of the TAGCAMS ground test results or substantially improved upon the ground test knowledge. In addition, the TAGCAMS calibration observations identified the source of a spacecraft outgassing feature that guided successful remediation efforts prior to asteroid arrival.
Full text
Available for:
DOBA, EMUNI, FIS, FZAB, GEOZS, GIS, IJS, IMTLJ, IZUM, KILJ, KISLJ, MFDPS, NLZOH, NUK, OBVAL, OILJ, PILJ, PNG, SAZU, SBCE, SBJE, SBMB, SBNM, UILJ, UKNU, UL, UM, UPUK, VKSCE, ZAGLJ
Hadal trenches are depocenters for organic material, and host intensified benthic microbial activity. The enhanced deposition is presumed to be reflected in elevated meiofaunal standing-stock, but ...available studies are ambiguous. Here, we investigate the distribution of meiofauna along the Atacama Trench axis and adjacent abyssal and bathyal settings in order to relate the meiofauna densities to proxies for food availability. Meiofauna densities peaked at the sediment surface and attenuated steeply with increasing sediment depth. The distribution mirrored the vertical profile of the microbial-driven oxygen consumption rate demonstrating a close linkage between microbial activity and meiofauna density. Meiofaunal standing-stock along the trench axis varied by a factor of two, but were markedly higher than values from the abyssal site at the oceanic plate. Overall, meiofaunal densities poorly correlated with common proxies for food availability such as total organic carbon and phytopigments, but strongly correlated with the microbial benthic O
consumption rate. We argue that microbial biomass likely represents an important meiofaunal food source for hadal meiofauna. Observations from three trench systems underlying surface water of highly different productivity confirmed elevated meiofaunal densities at the trench axis as compared to abyssal sites on oceanic plates. Food availability appear to drive elevated abundance and variations in meiofauna densities in hadal sediments.
Full text
Available for:
IZUM, KILJ, NUK, PILJ, PNG, SAZU, UL, UM, UPUK
PDF
