Purification procedure of APEH-3<i>_Ss</i> enzyme from <i>S. solfataricus</i>.

*<p>One unit of APEH activity (U) is defined as the amount of enzyme required to hydrolyze 1 µmol of Ac-L-<i>p</i>NA per min under the assay conditions.</p

Intestinal adverse outcomes in COVID-19: current evidence and uncertainties using the Adverse Outcome Pathway framework

Presentation to the European Society of Toxicology in Vitro (ESTIV) November 2022 Search for CCTE records in EPA’s Science Inventory by typing in the title at this link. https://cfpub.epa.gov/si/si_public_search_results.cfm?advSearch=true&showCriteria=2&keyword=CCTE&TIMSType=&TIMSSubTypeID=&epaNumber=&ombCat=Any&dateBeginPublishedPresented=07/01/2017&dateEndPublishedPresented=&dateBeginUpdated=&dateEndUpdated=&DEID=&personName=&personID=&role=Any&journalName=&journalID=&publisherName=&publisherID=&sortBy=pubDate&count=25</p

APEH structure and function analysis.

<p>(<b>A</b>) CD spectra performed at 80°C and 90°C for APEH<i>_Ss</i> or (<b>B</b>) APEH-3<i>_Ss</i>, respectively. (<b>C</b>) Effect of chaotropic agent on APEH<i>_Ss</i>, and (<b>D</b>) APEH-3<i>_Ss</i>: the enzymatic activity was measured using Ac-A-<i>p</i>NA for APEH-3<i>_Ss</i> and Ac-L-<i>p</i>NA for APEH<i>_Ss</i> as substrates, after addition of urea () or guanidine hydrochloride (); the activities were recovered after renaturation by extensive dialysis (); by comparison, mammalian APEH behavior <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037921#pone.0037921-Senthilkumar1" target="_blank">[<i>ref</i> 28]</a> is shown in <b>D </b><b><i>insert</i></b>. (<b>E</b>) Size exclusion chromatography elution profiles: native APEH-3<i>_Ss</i> (<b>^__</b>); APEH-3<i>_Ss</i> treated with GnHCl (<b>•••</b>); GnHCl-treated APEH-3<i>_Ss</i> after extensive dialysis (<b>---</b>).</p

Purification and sequence analysis of APEH-3<i>_Ss</i> from <i>Sulfolobus solfataricus</i>.

<p>(<b>A</b>) SDS-PAGE of purified APEH-3<i>_Ss</i>; lane 1, molecular weight markers (myosin 212.0 kDa, MBP-β-galactosidase 158.0 kDa, β-galactosidase 116.0 kDa, phosphorylase b 97.2 kDa, serum albumin 66.4 kDa, glutamic dehydrogenase 56.6 kDa, MBP2 <i>E. coli</i> 42.7 kDa, thioredoxin reductase 34.6 kDa and triosephosphate isomerase 27.0 kDa); lane 2, protein pattern of the cytoplasmic fraction obtained after cell lysis; lane 3, purified APEH-3<i>_Ss</i>; lane 4, purified APEH<i>_Ss</i>. (<b>B</b>) Amino acid sequence deduced from the APEH-3<i>_Ss</i>-coding gene (<i>sso2693</i>). The peptides identified by LC-MS/MS are highlighted in grey and the N-terminal amino acid sequence of the purified protease is boxed.</p

Phylogenetic analysis of APEH<i>_Ss</i> and APEH-3<i>_Ss</i> proteins in the archaeal kingdom.

<p>The cladogram shown in <b>A</b> (APEH<i>_Ss</i>) includes 43 archaeal organisms, while in the tree shown in <b>B</b> (APEH-3<i>_Ss</i>) only 15 species were retrieved, all belonging to <i>Crenarchaeota phylum</i>. The organisms in both trees are color coded according to the NCBI taxonomy. The trees were constructed by running Blast Explorer <a href="http://www.plosone.org/article/info:doi/10.1371/journal.pone.0037921#pone.0037921-Dereeper2" target="_blank">[41]</a>, in order to identify public similar sequences. For each run, proteins corresponding to hits pertaining to the Archaea, with BLAST e-value lower than 1E-20 and covering at least 50% of query sequence, were selected and fed to the “One Click Mode Phylogeny analysis” pipeline available on <a href="http://www.phylogeny.fr" target="_blank">www.phylogeny.fr</a>.</p

Transcriptional analysis under thermal stress.

<p>(<b>A–B</b>) Comparisons of <i>apeh_Ss</i> (squares) and <i>apeh-3_Ss</i> (triangles) fold changes of gene transcriptions under standard <b>()</b> or thermal stress <b>()</b> conditions, normalized respect to the expression level at t = t₀ of 23S gene.</p

Transcriptional analysis under oxidative stress.

<p>(<b>A–B</b>) Comparisons of <i>apeh_Ss</i> (squares) and <i>apeh-3_Ss</i> (triangles) fold changes of gene transcriptions under standard <b>()</b> or oxidative <b>()</b> conditions, normalized respect to the expression level at t = t₀ of 23S gene.</p

Towards Plant Species Identification in Complex Samples: A Bioinformatics Pipeline for the Identification of Novel Nuclear Barcode Candidates

<div><p>Monitoring of the food chain to fight fraud and protect consumer health relies on the availability of methods to correctly identify the species present in samples, for which DNA barcoding is a promising candidate. The nuclear genome is a rich potential source of barcode targets, but has been relatively unexploited until now. Here, we show the development and use of a bioinformatics pipeline that processes available genome sequences to automatically screen large numbers of input candidates, identifies novel nuclear barcode targets and designs associated primer pairs, according to a specific set of requirements. We applied this pipeline to identify novel barcodes for plant species, a kingdom for which the currently available solutions are known to be insufficient. We tested one of the identified primer pairs and show its capability to correctly identify the plant species in simple and complex samples, validating the output of our approach.</p></div

Development, Optimization, and Single Laboratory Validation of an Event-Specific Real-Time PCR Method for the Detection and Quantification of Golden Rice 2 Using a Novel Taxon-Specific Assay

In this study, we developed, optimized, and in-house validated a real-time PCR method for the event-specific detection and quantification of Golden Rice 2, a genetically modified rice with provitamin A in the grain. We optimized and evaluated the performance of the taxon (targeting rice Phospholipase D α2 gene)- and event (targeting the 3′ insert-to-plant DNA junction)-specific assays that compose the method as independent modules, using haploid genome equivalents as unit of measurement. We verified the specificity of the two real-time PCR assays and determined their dynamic range, limit of quantification, limit of detection, and robustness. We also confirmed that the taxon-specific DNA sequence is present in single copy in the rice genome and verified its stability of amplification across 132 rice varieties. A relative quantification experiment evidenced the correct performance of the two assays when used in combination

Bioinformatics pipeline for the identification of DNA barcodes in the nuclear genome.

<p>Scheme representing the flow of the different steps in the bioinformatics pipeline designed to process input sequences (top left) in order to output potential primer pairs amplifying novel DNA barcoding targets (bottom right). See text and Supporting Information for details.</p