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All sources NASA NTRS DOE OSTI

DOE OSTI2023

gRNA-SeqRET: a universal tool for targeted and genome-scale gRNA design and sequence extraction for prokaryotes and eukaryotes

High-throughput genetic screening is frequently employed to rapidly associate gene with phenotype and establish sequence-function relationships. With the advent of CRISPR technology, and the ability to functionally interrogate previously genetically recalcitrant organisms, non-model organisms can be investigated using pooled guide RNA (gRNA) libraries and sequencing-based assays to quantitatively assess fitness of every targeted locus in parallel. To aid the construction of pooled gRNA assemblies, we have developed an in silico design workflow for gRNA selection using the gRNA Sequence Region Extraction Tool (gRNA-SeqRET). Built upon the previously developed CCTop, gRNA-SeqRET enables automated, scalable design of gRNA libraries that target user-specified regions or whole genomes of any prokaryote or eukaryote. Additionally, gRNA-SeqRET automates the bulk extraction of any regions of sequence relative to genes or other features, aiding in the design of homology arms for insertion or deletion constructs. We also assess in silico the application of a designed gRNA library to other closely related genomes and demonstrate that for very closely related organisms Average Nucleotide Identity (ANI) > 95% a large fraction of the library may be of relevance. The gRNA-SeqRET web application pipeline can be accessed at https://grna.jgi.doe.gov. The source code is comprised of freely available software tools and customized Python scripts, and is available at https://bitbucket.org/berkeleylab/grnadesigner/src/master/ under a modified BSD open-source license (https://bitbucket.org/berkeleylab/grnadesigner).

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2023

Evaluation of Potential In Vitro Recombination Events in Codon Deoptimized FMDV Strains

Codon deoptimization (CD) has been recently used as a possible strategy to derive foot-and-mouth disease (FMD) live-attenuated vaccine (LAV) candidates containing DIVA markers. However, reversion to virulence, or loss of DIVA, from possible recombination with wild-type (WT) strains has yet to be analyzed. An in vitro assay was developed to quantitate the levels of recombination between WT and a prospective A24-P2P3 partially deoptimized LAV candidate. By using two genetically engineered non-infectious RNA templates, we demonstrate that recombination can occur within non-deoptimized viral genomic regions (i.e., 3'end of P3 region). The sequencing of single plaque recombinants revealed a variety of genome compositions, including full-length WT sequences at the consensus level and deoptimized sequences at the sub-consensus/consensus level within the 3'end of the P3 region. Notably, after further passage, two recombinants that contained deoptimized sequences evolved to WT. Overall, recombinants featuring large stretches of CD or DIVA markers were less fit than WT viruses. Our results indicate that the developed assay is a powerful tool to evaluate the recombination of FMDV genomes in vitro and should contribute to the improved design of FMDV codon deoptimized LAV candidates.

59 BASIC BIOLOGICAL SCIENCES↗

NASA NTRS1973

Data multiplexer using a tree switch

Self-decoding FET-hybrid or integrated-circuit tree configuration uses minimum number of components and can be sequenced by clock or computer. Redundancy features can readily be incorporated into tree configuration; as tree grows in size and more sensors are included, percentage of parts that will affect given percentage of sensors steadily decreases.

Easton, R. A.↗

NASA NTRS1973

Basalt stratigraphy of southern Mare Serenitatis

The major features of the stratigraphic and structural sequence are summarized for Mare Serenitatis. The dark oldest basalts include flows coextensive with Mare Tranquillitatis and also flows and pyroclastic deposits (dark mantles) that erupted from the edge of the Serenitatis basin. The basin sagged, possibly isostatically, as basalts of intermediate age were emplaced. Sagging had nearly ceased by the time the youngest flows were deposited. Compressive movements including thrusting followed, and small extensional fissures formed in late Copernican time. The revised stratigraphic sequence that was elaborated has implications for lunar stratigraphy that transcend the boundaries of Mare Serenitatis. Dark mantle deposits and the darkest maria have commonly been assumed in geologic mapping to be relatively youthful. These assumptions must now be reevaluated and perhaps discarded.

Howard, K. A.↗

NASA NTRS1980

Automation of the Mark 3 field system

The software and hardware components which will enable hands off operation are described. The operation of the field system begins with the scheduling of observations. An interactive program, SKED, provides displays of mutual visibility, automatic calculation of telescope slewing times, and the ability to list and edit the schedule. The output of SKED is a schedule file containing commands in the Standard Notation for Astronomy Procedures (SNAP) language which the field system uses for controlling events during the experiment. The most important features of SNAP include sophisticated time sequencing of events, automatic logging of all commands and responses, and the ability to define often used sequences of commands as procedures. A control program, BOSS, running in an HP 1000 minicomputer, reads the SNAP commands from a schedule file and interprets them in terms of commands and requests to devices. Interactive command input is possible through the operator's display terminal. Communication with all of the Mark 3 electronics modules is done via a small general purpose interface board (a microprocessor based ASCII transceiver) which has been installed in each module. Additional devices are controlled and monitored using the IEEE 488 General Purpose Interface Bus.

Vandenbere, N. R.↗

NASA NTRS1990

The evolution of Tharsis: Implications of gravity, topography, and tectonics

Dominating the Western Hemisphere of Mars, the Tharsis rise is an elongate area centered on Syria Planum that ascends as much as 8 to 10 km above the datum. It is intensely fractured by long, narrow grabens that extend radially hundreds of kilometers beyond the rise and is ringed by mostly concentric wrinkle ridges that formed over 2,000 km from the center of the rise. Its size, involving a full hemisphere of Mars, gives it a central role in the thermo-tectonic evolution of the planet and has stimulated a number of studies attempting to determine the sequence of events responsible for this feature. The constraints that gravity and topography data place on the current structure of Tharsis, along with insights into its development derived from comparisons of detailed regional mapping of faulting with theoretical deformation models are reviewed. Finally, a self-consistent model for the structure of Tharsis is proposed.

Banerdt, W. B.↗

NASA NTRS1996

Calcium-Dependent Protein Kinase Genes in Corn Roots

Two cDNAs encoding Ca-2(+) - Dependent Protein Kinases (CDPKs), Corn Root Protein Kinase 1 and 2 (CRPK 1, CRPK 2) were isolated from the root tip library of corn (Zea mays L., cv. Merit) and their nucleotide sequences were determined. Deduced amino acid sequences of both the clones have features characteristic of plant CDPKS, including all 11 conserved serine/threonine kinase subdomains, a junction domain and a calmodulin-like domain with four Ca-2(+), -binding sites. Northern analysis revealed that CRPKI mRNA is preferentially expressed in roots, especially in the root tip; whereas, the expression of CRPK2 mRNA was very low in all the tissues tested. In situ hybridization experiments revealed that CRPKI mRNA is highly expressed in the root apex, as compared to other parts of the root. Partially purified CDPK from the root tip phosphorylates syntide-2, a common peptide substrate for plant CDPKs, and the phosphorylation was stimulated 7-fold by the addition of Ca-2(+). Our results show that two CDPK isoforms are expressed in corn roots and they may be involved in the Ca-2(+)-dependent signal transduction process.

Takezawa, D.↗

NASA NTRS1998

The Marshall Automated Wind Algorithm for Geostationary Satellite Wind Applications

The Marshall Automated Wind (MAW) algorithm was developed over a decade ago in support of specialized studies of mesoscale meteorology. In recent years, the algorithm has been generalized to address global climate issues and other specific objectives related to NASA missions. The MAW algorithm uses a tracking scheme which minimizes image brightness temperature differences in a sequence of satellite images to determine feature displacement (winds). With the appropriate methodology accurate satellite derived winds can be obtained from visible, infrared, and water vapor imagery. Typical errors are less than 4 m/s but depend on the quality and control constraints used in post-processing. Key to this success is the judicious use of template size and search area used for tracking, image resolution and time sampling, and selection of appropriate statistical constraints which may vary with image type and desired application. The conference paper and subsequent poster will provide details of the technique and examples of its application.

Jedlovec, Gary J.↗

DOE OSTI2018

Multiple Laser Tracker Synchronization for the Vibration Analysis

Measuring the mechanical coupling and rigidity of a large apparatus presents unique challenges for 3D metrology. In this case the motion of well separated points attached to a rotor blade must be rapidly and repeatedly measured and the resulting data synchronized. This presentation describes the methodology and results for the mechanical coupling test of the magnetic field mapping system (FMS) for the Mu2e experiment at Fermi National Accelerator Laboratory (Fermilab). Three laser trackers, 2 API Radians and an API TIII, simultaneously measured the position of three cylindrical corner cube reflectors mounted on the FMS while the system was in motion and after coming to rest. The laser trackers took measurements at 83Hz for the T3 and 100Hz for the Radians, with fine spatial resolution, resulting in over 20,000 points measured per instrument in observation cycle. The laser tracker hardware and utilized software are unable to synchronize the three independent data streams directly during the measurement process. Instead a custom software program was developed to perform this synchronization of the data sequences during post-processing using features of the targets motion common to all three fiducial locations.

43 PARTICLE ACCELERATORS↗

DOE OSTI2019

Hanford Tank Side Cesium Removal System: Process Description and Performance Metrics - 19613

The Tank Side Cesium Removal (TSCR) System is a modular, quickly-deployable and passively safe system for efficiently removing solids and cesium from alkaline liquid waste raffinate created by spent fuel reprocessing on the Hanford site. It further describes TSCR component design features, system performance metrics and operational sequences used for liquid waste treatment, as well as spent ion exchange column (IXC) replacement and storage. This presentation should be of great interest to parties responsible for treatment and disposal of liquid waste containing high concentrations of cesium. (authors)

12 MANAGEMENT OF RADIOACTIVE AND NON-RADIOACTIVE W↗

NASA NTRS2004

Feature-Identification and Data-Compression Software

A report discusses the continuing development of Windows Interface for Nominal Displacement Selection (WINDS), a computer program for automated analysis of images of the Sun and planets acquired by scientific instruments aboard spacecraft. WINDS is intended to afford capabilities for identification of features, measurement of displacements and velocities, analysis of terrain and of atmospheres, and synthesis of animation sequences of images of terrains and atmospheres from small sets of samples by use of velocity-based interpolation. A major element of WINDS will be a nonlinear correlator capable of tracking small features in complex image sequences. For dynamic image sequences, the correlator will enable compression of data by factors >100. In processing image data, WINDS will take account of such factors as texture in image data, rotation of features during measurement intervals, effects of viewing and solar illumination angles, and vertical structures of atmospheres. WINDS will also take account of positions, aiming directions, and fields of view of cameras to determine three-dimensional feature structures by use of triangulation and stereoscopic analysis techniques.

Jong, Eric De↗

NASA NTRS1972

High resolution spectroscopy of the disk chromosphere. II - Time sequence observations of Ca II H and K emissions.

Two independent sets of high resolution time series spectra of the Ca II H and K emission obtained at the Solar Tower and at the Big Dome of the Sacramento Peak Observatory on September 11th, 1971 are reported. The evolutionary behavior of the emission first reported by Wilson and Evans is confirmed, but the detail of the evolution is found to be more complex. In one case, a doubly peaked feature showing some K3 emission evolves into a single K2 (red) peak with no K3 emission. Coincidentally, a neighboring doubly peaked feature evolves to a very strong blue peak. In an entirely independent sequence a doubly peaked feature evolves into a single red peak. The K2 emission then fades completely although the continuum threads are still strong. Finally a strong K2 blue peak appears. It is concluded that the observed evolution of the K2 emission is due to temporal variations in the physical conditions which give rise to them.

Wilson, P. R.↗

DOE OSTI2023

CPGminer: An Interactive Dashboard to Explore the Genomic Features and Taxonomy of Complete Prokaryotic Genomes

Prokaryotes, the earliest forms of life on Earth, play crucial roles in global biogeochemical processes in virtually all ecosystems. The ever-increasing amount of prokaryotic genome sequencing data provides a wealth of information to examine fundamental and applied questions through systematic genome comparison. Genomic features, such as genome size and GC content, and taxonomy-centric genomic features of complete prokaryotic genomes (CPGs) are crucial for various fields of microbial research and education, yet they are often overlooked. Additionally, creating systematically curated datasets that align with research concerns is an essential yet challenging task for wet-lab researchers. In this study, we introduce CPGminer, a user-friendly tool that allows researchers to quickly and easily examine the genomic features and taxonomy of CPGs and curate genome datasets. We also provide several examples to demonstrate its practical utility in addressing descriptive questions.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2026

Data for FUN-PROSE: A Deep Learning Approach to Predict Condition-Specific Gene Expression in Fungi

mRNA levels of all genes in a genome is a critical piece of information defining the overall state of the cell in a given environmental condition. Being able to reconstruct such condition-specific expression in fungal genomes is particularly important to metabolically engineer these organisms to produce desired chemicals in industrially scalable conditions. Most previous deep learning approaches focused on predicting the average expression levels of a gene based on its promoter sequence, ignoring its variation across different conditions. Here we present FUN-PROSE—a deep learning model trained to predict differential expression of individual genes across various conditions using their promoter sequences and expression levels of all transcription factors. We train and test our model on three fungal species and get the correlation between predicted and observed condition-specific gene expression as high as 0.85. We then interpret our model to extract promoter sequence motifs responsible for variable expression of individual genes. We also carried out input feature importance analysis to connect individual transcription factors to their gene targets. A sizeable fraction of both sequence motifs and TF-gene interactions learned by our model agree with previously known biological information, while the rest corresponds to either novel biological facts or indirect correlations.

Genomics↗

DOE OSTI2020

RNAcentral 2021: secondary structure integration, improved sequence search and new member databases

RNAcentral is a comprehensive database of non-coding RNA (ncRNA) sequences that provides a single access point to 44 RNA resources and >18 million ncRNA sequences from a wide range of organisms and RNA types. RNAcentral now also includes secondary (2D) structure information for >13 million sequences, making RNAcentral the world’s largest RNA 2D structure database. The 2D diagrams are displayed using R2DT, a new 2D structure visualization method that uses consistent, reproducible and recognizable layouts for related RNAs. The sequence similarity search has been updated with a faster interface featuring facets for filtering search results by RNA type, organism, source database or any keyword. This sequence search tool is available as a reusable web component, and has been integrated into several RNAcentral member databases, including Rfam, miRBase and snoDB. To allow for a more fine-grained assignment of RNA types and subtypes, all RNAcentral sequences have been annotated with Sequence Ontology terms. The RNAcentral database continues to grow and provide a central data resource for the RNA community. RNAcentral is freely available at https://rnacentral.org.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2018

Detecting Outliers in Network Transfers with Feature Extraction

Reliable file transfers are essential for successful science computations and experiments that need large complex data files to be moved across long distances. Networks use protocols such as TCP and UDP to support file transfers, and their performance degrades under packet losses and duplications, thereby adversely affecting science applications. Hence, scientists and engineers often monitor network statistics to find and repair network problems that cause such degradations.Outliers or anomalies can be detected using statistical and machine learning methods that highlight abnormal behaviors. However, without knowing what is normal, it is very difficult and subjective to determine the anomalies. In this paper, we investigate statistical and machine learning approaches to extract unknown or unsupervised features from TCP and perfSONAR data sets, with the aim to identify anomalies such as packet loss, duplication and retransmission sequencing that affect file transfer performance. Simple statistical and feature extraction methods (e.g. PCA) have shown that detecting anomalies is simpler if labeled data sets of normal behavior are available. While perfSONAR logs record losses,they do not detect duplications and reordering. We show that PCA applied to TCP statistics is able to determine abnormal behavior in the above cases. In particular, our results show that simple feature extraction techniques provide building blocks for more advanced network monitoring tools that can quickly flag anomalies using simple measurements, thereby helping to build confidence in file transfer reliability.

Rao, Nageswara S.↗

DOE OSTI2022

Classification of bacterial plasmid and chromosome derived sequences using machine learning

Plasmids are important genetic elements that facilitate horizonal gene transfer between bacteria and contribute to the spread of virulence and antimicrobial resistance. Most bacterial genome sequences in the public archives exist in draft form with many contigs, making it difficult to determine if a contig is of chromosomal or plasmid origin. Using a training set of contigs comprising 10,584 chromosomes and 10,654 plasmids from the PATRIC database, we evaluated several machine learning models including random forest, logistic regression, XGBoost, and a neural network for their ability to classify chromosomal and plasmid sequences using nucleotide k-mers as features. Based on the methods tested, a neural network model that used nucleotide 6-mers as features that was trained on randomly selected chromosomal and plasmid subsequences 5kb in length achieved the best performance, outperforming existing out-of-the-box methods, with an average accuracy of 89.38% ± 2.16% over a 10-fold cross validation. The model accuracy can be improved to 92.08% by using a voting strategy when classifying holdout sequences. In both plasmids and chromosomes, subsequences encoding functions involved in horizontal gene transfer—including hypothetical proteins, transporters, phage, mobile elements, and CRISPR elements—were most likely to be misclassified by the model. This study provides a straightforward approach for identifying plasmid-encoding sequences in short read assemblies without the need for sequence alignment-based tools.

59 BASIC BIOLOGICAL SCIENCES↗

DOE OSTI2020

*-DCC: A platform to collect, annotate, and explore a large variety of sequencing experiments

Background: Over the past few years the variety of experimental designs and protocols for sequencing experiments increased greatly. To ensure the wide usability of the produced data beyond an individual project, rich and systematic annotation of the underlying experiments is crucial. Findings: We first developed an annotation structure that captures the overall experimental design as well as the relevant details of the steps from the biological sample to the library preparation, the sequencing procedure, and the sequencing and processed files. Through various design features, such as controlled vocabularies and different field requirements, we ensured a high annotation quality, comparability, and ease of annotation. The structure can be easily adapted to a large variety of species. We then implemented the annotation strategy in a user-hosted web platform with data import, query, and export functionality. Conclusions: We present here an annotation structure and user-hosted platform for sequencing experiment data, suitable for lab-internal documentation, collaborations, and large-scale annotation efforts.

59 BASIC BIOLOGICAL SCIENCES↗