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At least 1,495 records · Page 83
All sourcesNASA NTRSDOE OSTI
DOE OSTI2022

Molecular mechanisms underlying host-induced gene silencing

Abstract Host-induced gene silencing (HIGS) refers to the silencing of genes in pathogens and pests by expressing homologous double-stranded RNAs (dsRNA) or artificial microRNAs (amiRNAs) in the host plant. The discovery of such trans-kingdom RNA silencing has enabled the development of RNA interference-based approaches for controlling diverse crop pathogens and pests. Although HIGS is a promising strategy, the mechanisms by which these regulatory RNAs translocate from plants to pathogens, and how they induce gene silencing in pathogens, are poorly understood. This lack of understanding has led to large variability in the efficacy of various HIGS treatments. This variability is likely due to multiple factors, such as the ability of the target pathogen or pest to take up and/or process RNA from the host, the specific genes and target sequences selected in the pathogen or pest for silencing, and where, when, and how the dsRNAs or amiRNAs are produced and translocated. In this review, we summarize what is currently known about the molecular mechanisms underlying HIGS, identify key unanswered questions, and explore strategies for improving the efficacy and reproducibility of HIGS treatments in the control of crop diseases.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

A method for achieving complete microbial genomes and improving bins from metagenomics data

Metagenomics facilitates the study of the genetic information from uncultured microbes and complex microbial communities. Assembling complete microbial genomes ( i.e ., circular with no misassemblies) from metagenomics data is difficult because most samples have high organismal complexity and strain diversity. Less than 100 circularized bacterial and archaeal genomes have been assembled from metagenomics data despite the thousands of datasets that are available. Circularized genomes are important for (1) building a reference collection as scaffolds for future assemblies, (2) providing complete gene content of a genome, (3) confirming little or no contamination of a genome, (4) studying the genomic context and synteny of genes, and (5) linking protein coding genes to ribosomal RNA genes to aid metabolic inference in 16S rRNA gene sequencing studies. We developed a method to achieve circularized genomes using iterative assembly, binning, and read mapping. In addition, this method exposes potential misassemblies from k-mer based assemblies. We chose species of the Candidate Phyla Radiation (CPR) to focus our initial efforts because they have small genomes and are only known to have one ribosomal RNA operon. We present 34 circular CPR genomes, one circular Margulisbacteria genome, and two circular megaphage genomes from 19 public and published datasets. We demonstrate findings that would likely be difficult without circularizing genomes, including that ribosomal genes are likely not operonic in the majority of CPR, and that some CPR harbor diverged forms of RNase P RNA. Code and a tutorial for this method is available at https://github.com/lmlui/Jorg .

Lui, Lauren↗
DOE OSTI2019

Radiation-damage investigation of a DNA 16-mer

In macromolecular crystallography, a great deal of effort has been invested in understanding radiation-damage progression. While the sensitivity of protein crystals has been well characterized, crystals of DNA and of DNA–protein complexes have not thus far been studied as thoroughly. Here, a systematic investigation of radiation damage to a crystal of a DNA 16-mer diffracting to 1.8 Å resolution and held at 100 K, up to an absorbed dose of 45 MGy, is reported. The RIDL ( Radiation-Induced Density Loss ) automated computational tool was used for electron-density analysis. Both the global and specific damage to the DNA crystal as a function of dose were monitored, following careful calibration of the X-ray flux and beam profile. The DNA crystal was found to be fairly radiation insensitive to both global and specific damage, with half of the initial diffraction intensity being lost at an absorbed average diffraction-weighted dose, D 1/2 , of 19 MGy, compared with 9 MGy for chicken egg-white lysozyme crystals under the same beam conditions but at the higher resolution of 1.4 Å. The coefficient of sensitivity of the DNA crystal was 0.014 Å 2 MGy -1 , which is similar to that observed for proteins. These results imply that the significantly greater radiation hardness of DNA and RNA compared with protein observed in a DNA–protein complex and an RNA–protein complex could be due to scavenging action by the protein, thereby protecting the DNA and RNA in these studies. In terms of specific damage, the regions of DNA that were found to be sensitive were those associated with some of the bound calcium ions sequestered from the crystallization buffer. In contrast, moieties farther from these sites showed only small changes even at higher doses.

Instruments & Instrumentation↗
DOE OSTI2015

Improved crystallization and diffraction of caffeine-induced death suppressor protein 1 (Cid1)

The use of truncation and RNA-binding mutations of caffeine induced death suppressor protein 1 (Cid1) as a means to enhance crystallogenesis leading to an improvement of X-ray diffraction resolution by 1.5 Å is reported. The post-transcriptional addition of uridines to the 3′-end of RNAs is an important regulatory process that is critical for coding and noncoding RNA stability. In fission yeast and metazoans this untemplated 3′-uridylylation is catalysed by a single family of terminal uridylyltransferases (TUTs) whose members are adapted to specific RNA targets. In Schizosaccharomyces pombe the TUT Cid1 is responsible for the uridylylation of polyadenylated mRNAs, targeting them for destruction. In metazoans, the Cid1 orthologues ZCCHC6 and ZCCHC11 uridylate histone mRNAs, targeting them for degradation, but also uridylate microRNAs, altering their maturation. Cid1 has been studied as a model TUT that has provided insights into the larger and more complex metazoan enzyme system. In this paper, two strategies are described that led to improvements both in the crystallogenesis of Cid1 and in the resolution of diffraction by ∼1.5 Å. These advances have allowed high-resolution crystallo@@graphic studies of this TUT system to be initiated.

75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND↗
DOE OSTI2015

The structure of a contact-dependent growth-inhibition (CDI) immunity protein from Neisseria meningitidis MC58

Contact-dependent growth inhibition (CDI) is an important mechanism of intercellular competition between neighboring Gram-negative bacteria. CDI systems encode large surface-exposed CdiA effector proteins that carry a variety of C-terminal toxin domains (CdiA-CTs). All CDI + bacteria also produce CdiI immunity proteins that specifically bind to the cognate CdiA-CT and neutralize its toxin activity to prevent auto-inhibition. Here, the X-ray crystal structure of a CdiI immunity protein from Neisseria meningitidis MC58 is presented at 1.45 Å resolution. The CdiI protein has structural homology to the Whirly family of RNA-binding proteins, but appears to lack the characteristic nucleic acid-binding motif of this family. Sequence homology suggests that the cognate CdiA-CT is related to the eukaryotic EndoU family of RNA-processing enzymes. A homology model is presented of the CdiA-CT based on the structure of the XendoU nuclease from Xenopus laevis . Molecular-docking simulations predict that the CdiA-CT toxin active site is occluded upon binding to the CdiI immunity protein. Together, these observations suggest that the immunity protein neutralizes toxin activity by preventing access to RNA substrates.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2015

Structural characterization of the principal mRNA-export factor Mex67–Mtr2 from Chaetomium thermophilum

The crystal structures of the individual domains of the Mex67–Mtr2 complex from C. thermophilum have been determined and their arrangement in solution has been studied by SAXS. Members of the Mex67–Mtr2/NXF–NXT1 family are the principal mediators of the nuclear export of mRNA. Mex67/NXF1 has a modular structure based on four domains (RRM, LRR, NTF2-like and UBA) that are thought to be present across species, although the level of sequence conservation between organisms, especially in lower eukaryotes, is low. Here, the crystal structures of these domains from the thermophilic fungus Chaetomium thermophilum are presented together with small-angle X-ray scattering (SAXS) and in vitro RNA-binding data that indicate that, not withstanding the limited sequence conservation between different NXF family members, the molecules retain similar structural and RNA-binding properties. Moreover, the resolution of crystal structures obtained with the C. thermophilum domains was often higher than that obtained previously and, when combined with solution and biochemical studies, provided insight into the structural organization, self-association and RNA-binding properties of Mex67–Mtr2 that facilitate mRNA nuclear export.

75 CONDENSED MATTER PHYSICS, SUPERCONDUCTIVITY AND↗
DOE OSTI2018

Crystal structure of the ribonuclease-P-protein subunit from Staphylococcus aureus

Staphylococcus aureus ribonuclease-P-protein subunit (RnpA) is a promising antimicrobial target that is a key protein component for two essential cellular processes, RNA degradation and transfer-RNA (tRNA) maturation. The first crystal structure of RnpA from the pathogenic bacterial species, S. aureus , is reported at 2.0 Å resolution. The structure presented maintains key similarities with previously reported RnpA structures from bacteria and archaea, including the highly conserved RNR-box region and aromatic residues in the precursor-tRNA 5'-leader-binding domain. This structure will be instrumental in the pursuit of structure-based designed inhibitors targeting RnpA-mediated RNA processing as a novel therapeutic approach for treating S. aureus infections.

Ha, Lisha↗
DOE OSTI2016

Hepatitis C virus dynamics and cellular gene expression in uPA-SCID chimeric mice with humanized livers during intravenous silibinin monotherapy

Legalon SIL (SIL) is a chemically hydrophilized version of silibinin, an extract of milk thistle ( Silybum marianum ) seeds that has exhibited hepatoprotective and antiviral effectiveness against hepatitis C virus (HCV) in patients leading to viral clearance in combination with ribavirin. In this paper, to elucidate the incompletely understood mode of action of SIL against HCV, mathematical modelling of HCV kinetics and human hepatocyte gene expression studies were performed in uPA-SCID-chimeric mice with humanized livers. Chronically HCV-infected mice (n = 15) were treated for 14 days with daily intravenous SIL at 469, 265 or 61.5 mg/kg. Serum HCV and human albumin (hAlb) were measured frequently, and liver HCV RNA was analysed at days 3 and 14. Microarray analysis of human hepatocyte gene expression was performed at days 0, 3 and 14 of treatment. While hAlb remained constant, a biphasic viral decline in serum was observed consisting of a rapid 1st phase followed by a second slower phase (or plateau with the two lower SIL dosings). SIL effectiveness in blocking viral production was similar among dosing groups (median ε = 77%). However, the rate of HCV-infected hepatocyte decline, δ, was dose-dependent. Intracellular HCV RNA levels correlated (r = 0.66, P = 0.01) with serum HCV RNA. Pathway analysis revealed increased anti-inflammatory and antiproliferative gene expression in human hepatocytes in SIL-treated mice. Finally, the results suggest that SIL could lead to a continuous second-phase viral decline, that is potentially viral clearance, in the absence of adaptive immune response along with increased anti-inflammatory and antiproliferative gene expression in human hepatocytes.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2020

2.7 Å cryo-EM structure of rotavirus core protein VP3, a unique capping machine with a helicase activity

In many viruses, including rotavirus (RV), the major pathogen of infantile gastroenteritis, capping of viral messenger RNAs is a pivotal step for efficient translation of the viral genome. In RV, VP3 caps the nascent transcripts synthesized from the genomic dsRNA segments by the RV polymerase VP1 within the particle core. Here, from cryo–electron microscopy, x-ray crystallography, and biochemical analyses, we show that VP3 forms a stable tetrameric assembly with each subunit having a modular domain organization, which uniquely integrates five distinct enzymatic steps required for capping the transcripts. In addition to the previously known guanylyl- and methyltransferase activities, we show that VP3 exhibits hitherto unsuspected RNA triphosphatase activity necessary for initiating transcript capping and RNA helicase activity likely required for separating the RNA duplex formed transiently during endogenous transcription. From our studies, we propose a new mechanism for how VP3 inside the virion core caps the nascent transcripts exiting from the polymerase.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2021

The structural plasticity of nucleic acid duplexes revealed by WAXS and MD

Double-stranded DNA (dsDNA) and RNA (dsRNA) helices display an unusual structural diversity. Some structural variations are linked to sequence and may serve as signaling units for protein-binding partners. Therefore, elucidating the mechanisms and factors that modulate these variations is of fundamental importance. While the structural diversity of dsDNA has been extensively studied, similar studies have not been performed for dsRNA. Because of the increasing awareness of RNA’s diverse biological roles, such studies are timely and increasingly important. We integrate solution x-ray scattering at wide angles (WAXS) with all-atom molecular dynamics simulations to explore the conformational ensemble of duplex topologies for different sequences and salt conditions. These tightly coordinated studies identify robust correlations between features in the WAXS profiles and duplex geometry and enable atomic-level insights into the structural diversity of DNA and RNA duplexes. Notably, dsRNA displays a marked sensitivity to the valence and identity of its associated cations.

99 GENERAL AND MISCELLANEOUS↗
DOE OSTI2018

Programmed DNA destruction by miniature CRISPR-Cas14 enzymes

CRISPR-Cas systems provide microbes with adaptive immunity to infectious nucleic acids and are widely employed as genome editing tools. These tools use RNA-guided Cas proteins whose large size (950 to 1400 amino acids) has been considered essential to their specific DNA- or RNA-targeting activities. Here we present a set of CRISPR-Cas systems from uncultivated archaea that contain Cas14, a family of exceptionally compact RNA-guided nucleases (400 to 700 amino acids). Despite their small size, Cas14 proteins are capable of targeted single-stranded DNA (ssDNA) cleavage without restrictive sequence requirements. Moreover, target recognition by Cas14 triggers nonspecific cutting of ssDNA molecules, an activity that enables high-fidelity single-nucleotide polymorphism genotyping (Cas14-DETECTR). Metagenomic data show that multiple CRISPR-Cas14 systems evolved independently and suggest a potential evolutionary origin of single-effector CRISPR-based adaptive immunity.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2018

Microbial rRNA Synthesis and Growth Compared through Quantitative Stable Isotope Probing with H 2 18 O

Growing bacteria have a high concentration of ribosomes to ensure sufficient protein synthesis, which is necessary for genome replication and cellular division. To elucidate whether metabolic activity of soil microorganisms is coupled with growth, we investigated the relationship between rRNA and DNA synthesis in a soil bacterial community using quantitative stable isotope probing (qSIP) with H 2 18 O. Most soil bacterial taxa were metabolically active and grew, and there was no significant difference between the isotopic composition of DNA and RNA extracted from soil incubated with H 2 18 O. The positive correlation between 18 O content of DNA and rRNA of taxa, with a slope statistically indistinguishable from 1 (slope = 0.96; 95% confidence interval [CI], 0.90 to 1.02), indicated that few taxa made new rRNA without synthesizing new DNA. There was no correlation between rRNA-to-DNA ratios obtained from sequencing libraries and the atom percent excess (APE) 18 O values of DNA or rRNA, suggesting that the ratio of rRNA to DNA is a poor indicator of microbial growth or rRNA synthesis. Our results support the notion that metabolic activity is strongly coupled to cellular division and suggest that nondividing taxa do not dominate soil metabolic activity. IMPORTANCE: Utilizing quantitative stable isotope probing of microbial RNA and DNA with H 2 18 O, we show that most soil taxa are metabolically active and grow because their nucleic acids are significantly labeled with 18 O. A majority of the populations that make new rRNA also grow, which argues against the common paradigm that most soil taxa are dormant. Moreover, our results indicate that relative sequence abundance-based RNA-to-DNA ratios, which are frequently used for identifying active microbial populations in the environment, underestimate the number of metabolically active taxa within soil microbial communities.

environmental microbiology↗
DOE OSTI2017

Corexit 9500 Enhances Oil Biodegradation and Changes Active Bacterial Community Structure of Oil-Enriched Microcosms

To better understand the impacts of Corexit 9500 on the structure and activity levels of hydrocarbon-degrading microbial communities, we analyzed next-generation 16S rRNA gene sequencing libraries of hydrocarbon enrichments grown at 5 and 25°C using both DNA and RNA extracts as the sequencing templates. Oil biodegradation patterns in both 5 and 25°C enrichments were consistent with those reported in the literature (i.e., aliphatics were degraded faster than aromatics). Slight increases in biodegradation were observed in the presence of Corexit at both temperatures. Differences in community structure were observed between treatment conditions in the DNA-based libraries. The 25°C consortia were dominated by Vibrio , Idiomarina , Marinobacter , Alcanivorax , and Thalassospira species, while the 5°C consortia were dominated by several species of the genera Flavobacterium , Alcanivorax , and Oleispira . Most of these genera have been linked to hydrocarbon degradation and have been observed after oil spills. Colwellia and Cycloclasticus , known aromatic degraders, were also found in these enrichments. The addition of Corexit did not have an effect on the active bacterial community structure of the 5°C consortia, while at 25°C, a decrease in the relative abundance of Marinobacter was observed. At 25°C, Thalassospira , Marinobacter , and Idiomarina were present at higher relative abundances in the RNA than DNA libraries, suggesting that they were active in degradation. Similarly, Oleispira was greatly stimulated by the addition of oil at 5°C While dispersants such as Corexit 9500 can be used to treat oil spills, there is still debate on the effectiveness on enhancing oil biodegradation and its potential toxic effect on oil-degrading microbial communities. The results of this study provide some insights on the microbial dynamics of hydrocarbon-degrading bacterial populations in the presence of Corexit 9500. Operational taxonomic unit (OTU) analyses indicated that several OTUs were inhibited by the addition of Corexit. Conversely, a number of OTUs were stimulated by the addition of the dispersant, many of which were identified as known hydrocarbon-degrading bacteria. The results highlight the value of using RNA-based methods to further understand the impact of dispersant on the overall activity of different hydrocarbon-degrading bacterial groups.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

Experimental Evolution of Extreme Resistance to Ionizing Radiation in Escherichia coli after 50 Cycles of Selection

In prior work (D. R. Harris et al., J Bacteriol 191:5240–5252, 2009, https://doi.org/10.1128/JB.00502-09; B. T. Byrne et al., Elife 3:e01322, 2014, https://doi.org/10.7554/eLife.01322), we demonstrated that Escherichia coli could acquire substantial levels of resistance to ionizing radiation (IR) via directed evolution. Major phenotypic contributions involved adaptation of organic systems for DNA repair. We have currently undertaken an extended effort to generate E. coli populations that are as resistant to IR as Deinococcus radiodurans . After an initial 50 cycles of selection using high-energy electron beam IR, four replicate populations exhibit major increases in IR resistance but have not yet reached IR resistance equivalent to D. radiodurans . Regular deep sequencing reveals complex evolutionary patterns with abundant clonal interference. Prominent IR resistance mechanisms involve novel adaptations to DNA repair systems and alterations in RNA polymerase. Adaptation is highly specialized to resist IR exposure, since isolates from the evolved populations exhibit highly variable patterns of resistance to other forms of DNA damage. Sequenced isolates from the populations possess between 184 and 280 mutations. IR resistance in one isolate, IR9-50-1, is derived largely from four novel mutations affecting DNA and RNA metabolism: RecD A90E, RecN K429Q, and RpoB S72N/RpoC K1172I. Additional mechanisms of IR resistance are evident. IMPORTANCE Some bacterial species exhibit astonishing resistance to ionizing radiation, with Deinococcus radiodurans being the archetype. As natural IR sources rarely exceed mGy levels, the capacity of Deinococcus to survive 5,000 Gy has been attributed to desiccation resistance. To understand the molecular basis of true extreme IR resistance, we are using experimental evolution to generate strains of Escherichia coli with IR resistance levels comparable to Deinococcus . Experimental evolution has previously generated moderate radioresistance for multiple bacterial species. Yet, these efforts could not take advantage of modern genomic sequencing technologies. In this report, we examine four replicate bacterial populations after 50 selection cycles. Genomic sequencing allows us to follow the genesis of mutations in populations throughout selection. Novel mutations affecting genes encoding DNA repair proteins and RNA polymerase enhance radioresistance. However, more contributors are apparent.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2016

Structural, Mechanistic, and Antigenic Characterization of the Human Astrovirus Capsid

Human astroviruses (HAstVs) are nonenveloped, positive-sense, single-stranded RNA viruses that are a leading cause of viral gastroenteritis. HAstV particles display T=3 icosahedral symmetry formed by 180 copies of the capsid protein (CP), which undergoes proteolytic maturation to generate infectious HAstV particles. Little is known about the molecular features that govern HAstV particle assembly, maturation, infectivity, and immunogenicity. Here we report the crystal structures of the two main structural domains of the HAstV CP: the core domain at 2.60-Å resolution and the spike domain at 0.95-Å resolution. Fitting of these structures into the previously determined 25-Å-resolution electron cryomicroscopy density maps of HAstV allowed us to characterize the molecular features on the surfaces of immature and mature T=3 HAstV particles. The highly electropositive inner surface of HAstV supports a model in which interaction of the HAstV CP core with viral RNA is a driving force in T=3 HAstV particle formation. Additionally, mapping of conserved residues onto the HAstV CP core and spike domains in the context of the immature and mature HAstV particles revealed dramatic changes to the exposure of conserved residues during virus maturation. Indeed, we show that antibodies raised against mature HAstV have reactivity to both the HAstV CP core and spike domains, revealing for the first time that the CP core domain is antigenic. Together, these data provide new molecular insights into HAstV that have practical applications for the development of vaccines and antiviral therapies. Astroviruses are a leading cause of viral diarrhea in young children, immunocompromised individuals, and the elderly. Despite the prevalence of astroviruses, little is known at the molecular level about how the astrovirus particle assembles and is converted into an infectious, mature virus. In this paper, we describe the high-resolution structures of the two main astrovirus capsid proteins. Fitting these structures into previously determined low-resolution maps of astrovirus allowed us to characterize the molecular surfaces of immature and mature astroviruses. Our studies provide the first evidence that astroviruses undergo viral RNA-dependent assembly. We also provide new insight into the molecular mechanisms that lead to astrovirus maturation and infectivity. Finally, we show that both capsid proteins contribute to the adaptive immune response against astrovirus. Together, these studies will help to guide the development of vaccines and antiviral drugs targeting astrovirus.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2019

Sudan Ebolavirus VP35-NP Crystal Structure Reveals a Potential Target for Pan-Filovirus Treatment

The filoviruses are etiological agents of life-threatening hemorrhagic fever with high mortality rate and risk of potential outbreak. Among members of this family, the Ebola (EBOV), Sudan (SUDV), and Marburg (MARV) viruses are considered the most pathogenic for humans. The ebolavirus nucleoprotein (NP) is the most abundant protein in infected cells and is essential for viral transcription and replication; thus, it represents an attractive target for therapeutic intervention. Here, we present the structure of SUDV NP in complex with the amino-terminal portion of the phosphoprotein VP35 at 2.3 Å. This structure captures VP35 chaperoning SUDV NP in a monomeric and RNA-free state. This transient state has been proposed to be key to maintaining a pool of monomeric and RNA-free NPs prior to NP-NP polymerization and encapsidation of the viral RNA genome. This structure also reveals a newly visualized interaction between NP and VP35, a well-defined beta sheet that is not present in previous structures. Affinity binding assays demonstrate that this beta sheet is essential for maintaining the high-affinity interaction between VP35 and a hydrophobic pocket on SUDV NP, and electron microscopy indicates the importance of this binding interaction to the oligomeric state and assembly of NP in human cells. Complementary structure-directed mutagenesis identifies critical residues conserved across the filovirus family that could be targeted by broadly effective antivirals.

59 BASIC BIOLOGICAL SCIENCES↗
DOE OSTI2025

divergence in basal transcriptomes of "Wild Type" Chlamydomonas reinhardtii strains

This is a study to compare the basal transcriptomes of several widely used laboratory strains of the Chlorophyte alga, Chlamydomonas reinhardtii. Given that there is a high degree of genetic diversity among the closely-related laboratory strains, we wished to examine how much variation there is at the transcriptome level. A panel of WT strains (CC-124, CC-125, CC-1009, CC-1690, CC-1691), all believed to be descended from a single zygospore isolated in 1945, were chosen based on their representing the oldest lineages among the standard laboratory strains. Additionally, CC-4532, which was the source for the current (v6) reference assembly, and CC-4533, which is the initial parental strain of the CLiP library collection of mutant strains, were also included in this study based on their significance to the Chlamydomonas community. All strains were grown in liquid cultures under identical, mixotrophic conditions (light + acetate) to mid-log phase before collecting mRNA for RNA-Seq analysis. Overall design: Pre-cultures of each strain (CC-124, CC-125, CC-1009, CC-1690, CC-1691, CC-4532, and CC-4533) were used to innoculate flasks of Tris-Acetate-Phosphate (TAP) media supplemented with Kropat's trace metals at a concentration of 1.5 x 10^4 cells/mL. Flasks were placed on a shaking platform at 180 RPM under 50-70 µmol x m^-2 x s^-1 of light. Cultures were grown until they reached ~2-3 x 10^6 cells/mL, about three days, before total RNA was collected and subjected RNA-Seq analysis.

Source record↗
DOE OSTI2019

P Finder

P Finder is written in C# and has been tested in multiple 64-bit Windows environments (Windows 7/8/Vista). It has a user-friendly graphical user interface (GUI) that allows for intuitive RNase P RNA identification and/or annotation. The user can select from four search criteria including the use multiple threads. In addition, the user can determine what display data they wish to incorporate in the output such as: bitscore, E-value, percent ID, start location, end location, the strand in which the sequence is located, and of course the sequence itself. P Finder is the only available algorithm that is capable of not only identifying the RNase P RNA for annotation but also differentiating among the five different RNase P RNA structural types (A, B, C, M, and T) with the option to select these data types for display in the final output.

Ellis, JosephC↗