Final Library Research Articles

Pin1 WW Domain Ligand Library Synthesized with an Easy Solid-Phase Phosphorylating Reagent.

Cell cycle regulatory enzyme Pin1 both catalyzes pSer/Thr-cis/trans-Pro isomerization and binds the same motif separately in its WW domain. To better understand the function of Pin1, a way to separate these activities is needed. An unnatural peptide library, R1CO-pSer-Pro-NHR2, was designed to identify ligands specific for the Pin1 WW domain. A new solid-phase phosphorylating reagent (SPPR) containing a phosphoramidite functional group was synthesized in one step from Wang resin. The SPPR was used in the preparation of the library by parallel synthesis. The final 315-member library was screened with our WW-domain-specific, enzyme-linked enzyme-binding assay (ELEBA). Four of the best hits were resynthesized, and the competitive dissociation constants were measured by ELEBA. NMR chemical-shift perturbations (CSP) of ligands with 15N-labeled Pin1 were used to measure Kd for the best four ligands directly, demonstrating that they were specific Pin1 WW domain ligands. Models of the ligands bound to the Pin1 WW domain were used to visualize the mode of binding in the WW domain.

Generation of a Phage Display Chicken Single-Chain Variable Fragment Library.

Phage-displayed antibody fragment libraries can be constructed using essentially any species that is easily immunized, as long as the immunoglobulin variable region gene sequences are known. This protocol describes the procedures for the generation of a phage-displayed chicken single-chain variable fragment (scFv) library after immunization with a target antigen. Briefly, the rearranged heavy chain variable region (V H ) genes and the λ light chain variable region (V λ ) genes are amplified separately and are linked through two separate PCR steps to give the final scFv genes. The genes are then cloned into pComb3XSS to generate the phage display chicken scFv library, which can then be used for test and final library ligations.

Highly pure DNA-encoded chemical libraries by dual-linker solid-phase synthesis.

The first drugs discovered using DNA-encoded chemical library (DEL) screens have entered late-stage clinical development. However, DEL technology as a whole still suffers from poor chemical purity resulting in suboptimal performance. In this work, we report a technique to overcome this issue through self-purifying release of the DEL after magnetic bead-based synthesis. Both the first and last building blocks of each assembled library member were linked to the beads by tethers that could be cleaved by mutually orthogonal chemistry. Sequential cleavage of the first and last tether, with washing in between, ensured that the final library comprises only the fully complete compounds. The outstanding purity attained by this approach enables a direct correlation of chemical display and encoding, allows for an increased chemical reaction scope, and facilitates the use of more diversity elements while achieving greatly improved signal-to-noise ratios in selections.

Design and implementation of an interactive virtual library based on its physical counterpart

The rapid technological advancements and the widespread adoption of the internet have diminished the role of the physical library as a main information resource. As the Metaverse is evolving, a revolutionary change is anticipated in how social relationships are perceived, within an educational context. It is therefore necessary for libraries to upgrade the services they provide to keep in line with the technological trends and be a part of this virtual revolution. It is believed that the design and development of a Virtual Reality (VR) library can be the community and knowledge hub the society needs. In this paper, the process of creating a partially digital replica of the Limassol Municipal University Library, a landmark for the city of Limassol, is examined by using photogrammetry and 3D modelling. A 3D platform was developed, where users have the perception that they are experiencing the actual library. To that end, a perceptual study was conducted, to understand the current usage of physical libraries, examine the users’ experience in VR, and identify the requirements and expectations in the development of a virtual library counterpart. Following the suggestions and observations from the perceptual study, five key scenarios were implemented that demonstrate the potential use of a virtual library. This work incorporates the fundamental VR attributes, such as immersiveness, realism, user interactivity and feedback as well as other features, such as animated NPCs, 3D audio, ray-casting and GUIs, that significantly augment the overall VR library user experience, presence as well as navigation autonomy. The main effort of this project was to produce a VR representation of an existing physical library, integrated with its key services, as a proof-of-concept, with emphasis on easy 24/7 access, functionality, and interactivity. The above attributes differentiate this work from existing studies. A detailed user evaluation study was conducted upon completion of the final VR library implementation, which firmly confirmed all its key attributes and future viability.

Leveraging the fundamentals of heat transfer and fluid mechanics in microscale geometries for automated next-generation sequencing library preparation

Next-generation sequencing (NGS) is emerging as a powerful tool for molecular diagnostics but remains limited by cumbersome and inefficient sample preparation. We present an innovative automated NGS library preparation system with a simplified mechanical design that exploits both macro- and microfluidic properties for optimizing heat transfer, reaction kinetics, mass transfer, fluid mechanics, adsorption–desorption rates, and molecular thermodynamics. Our approach introduces a unique two-cannula cylindrical capillary system connected to a programmable syringe pump and a Peltier heating element able to execute all steps with high efficiency. Automatic reagent movement, mixing, and magnetic bead-based washing with capillary-based thermal cycling (capillary-PCR) are completely integrated into a single platform. The manual 3-h library preparation process is reduced to less than 15 min of hands-on time via optimally pre-plated reagent plates, followed by less than 6 h of instrument run time during which no user interaction is required. We applied this method to two library preparation assays with different DNA fragmentation requirements (mechanical vs. enzymatic fragmentation), sufficiently limiting consumable use to one cartridge and one 384 well-plate per run. Our platform successfully prepared eight libraries in parallel, generating sequencing data for both human and Escherichia coli DNA libraries with negligible coverage bias compared to positive controls. All sequencing data from our libraries attained Phred (Q) scores > 30, mapping to reference genomes at 99% confidence. The method achieved final library concentrations and size distributions comparable with the conventional manual approach, demonstrating compatibility with downstream sequencing and subsequent data analysis. Our engineering design offers repeatability and consistency in the quality of sequence-able libraries, asserting the importance of mechanical design considerations that employ and optimize fundamental fluid mechanics and heat transfer properties. Furthermore in this work, we provide unique insights into the mechanisms of sample loss within NGS library preparation assays compared with automated adaptations and pinpoint areas in which the principles of thermodynamics, fluid mechanics, and heat transfer can improve future mechanical design iterations.

Abstract LB432: Identification of novel drug targets using the ADAPT biotargeting system on FFPE tissue from patients with glioblastoma multiforme

Abstract Glioblastoma Multiforme (GBM) is one of the most aggressive and lethal brain tumor types with a five-year survival rate of 6.9%. In 2023, more than 14,490 Americans were expected to receive a diagnosis. Although great progress has been achieved in protein target identification and treatment over the last few years, current treatment options are limited, and tumor recurrence occurs at a high rate due to treatment resistance. Aptamers, short single-stranded oligonucleotide ligands with high-affinity binding to molecular targets, are well suited tools for exploring novel targets in highly heterogeneous tumors with complex etiologies like GBM, where clinically relevant biomarkers are limited. In this study, aptamer-based cancer-specific drug target identification was performed by Adaptive Dynamic Artificial Polyligand Targeting (ADAPT) Biotargeting System. In short, a GBM-specific library was enriched through a Systematic Evolution of Ligands by EXponential enrichment (SELEX) process using micro dissected Formalin-Fixed Paraffin-Embedded (FFPE) tissues from GBM patients, along with whole body normal tissue for counterselection. Microdissection of tissue allows the partitioning of different types of cells to focus on specific cells of interest. The enriched libraries have been evaluated on clinical GBM and normal brain control samples through aptamer mediated tissue staining, resulting in preferential binding of enriched libraries compared to controls in the vast majority of GBM patients with little binding to normal brain samples. The final library was subsequently applied in an in-house developed library-based affinity pull-down assay using lysates from FFPE tissue of the same group of GBM patients and normal brain samples, followed by unbiased Mass Spectrometry for protein target identification. Combining the results from this proteomics analysis and data from the internal Whole Transcriptome Sequencing (WTS) database, we were able to identify several targets that are highly expressed in GBM and other cancer lineages. Antibody staining provided further validation of the expression levels and the cellular localization. The targets identified in this study include not only potential novel therapeutic targets, but also well-studied biomarkers and targets with available drugs already in clinical trials, demonstrating the platform has great potential for identifying high affinity binding and selectivity compared to traditional proteomics and other methods in drug target discovery. Citation Format: Xixi Wei, Myunggi An, Radhika Santhanam, Barbara Richardson, Lucas Vu, Justin Saul, Anna Walton, Michael Gee, Daniel Martin, Amanda Hubbard, Alexis Rodin, Matthew Rosenow, Tassilo Hornung. Identification of novel drug targets using the ADAPT biotargeting system on FFPE tissue from patients with glioblastoma multiforme [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 2 (Late-Breaking, Clinical Trial, and Invited Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(7_Suppl):Abstract nr LB432.

Use of Social Media by Library and Information Science Students

This study examined the use of social media by final year Library and Information Science students of Nnamdi Azikiwe University, Awka. Three research questions guided the study. The study employed descriptive survey design using questionnaire to gather information. The sample size is one hundred and sixty-five (165) undergraduate LIS final year students in the Department of Library and Information Science, Nnamdi Azikiwe University, Awka and Delta State University, Abraka. The data collected was analysed using mean and standard deviation to answer the research questions. The study findings revealed that the most used social media by final year LIS students are Facebook, WhatsApp, Instagram, YouTube and telegram while zoom, twitter, skype, myspace, blog and LinkedIn are least used. The final year LIS students used social media for different purposes such as; to obtain information for project writing, assignment, to supplement lecture notes, interacting with course mates/friends, socialization/entertainment and interaction with family members. The LIS students encountered various problems with the use of social media which include time management, distraction, addiction, encroachment on academic activities, high cost of internet connectivity and poor social media skills. The study recommended that the challenges of using social media should be tackle, so that students can properly use social media for information access.

Abstract 6255: Personalized medicine for DCIS

Abstract Background: In 2023, 55,720 women will be diagnosed with non-invasive ductal carcinoma in situ (DCIS) in the United States. We evaluated RNA sequencing (RNA-Seq), clinical and social factors that drive subsequent breast events in a DCIS study population enriched for Black women (30%). Methods: RNA was extracted from macrosections of archival formalin-fixed paraffin-embedded (FFPE) DCIS tissue from Resource Archival Human Breast Tissue (RAHBT) from the St. Louis Breast Tissue Registry at Washington University School of Medicine and the Dana Farber Cancer Institute patient cohorts. Tissues were prepared by microsection of 5 uM whole section from unstained slides (Fisher). RNA was extracted using Qiagen RNeasy FFPE Kits from macrosectioned DCIS specimens. Purified total RNA was evaluated for quality and quantity using the Agilent Fragment Analyzer RNA assay. Bulk RNA sequencing was piloted in DCIS case-control samples using the SMART-seq and Hybrid Capture platforms at the DFCI Molecular Biology Core Facilities. Raw reads were aligned to the GRCh38/hg38 reference genome using STAR. Preliminary quality control analysis of a curated gene list showed higher quality data using the Hybrid Capture technology. Libraries for Hybrid Capture were pooled in 8plex reactions and hybridized to Twist Exome 2.0 probes for target enrichment. The final library pools were sequenced on an Illumina NovaSeq. Cases were defined as women diagnosed with DCIS in 1999 or later with subsequent invasive breast cancer, local DCIS recurrence, or lobular CIS recurrence. Controls were defined as women diagnosed with DCIS in 1999 or later without subsequent breast events during a similar follow-up time. The R package DESeq2 was used for data analysis. Results: The mean age at DCIS diagnosis was 56.3 (Std Dev 10.23). The mean follow-up time was 147.8 months. Hybrid Capture RNA-Seq files are stored and analyzed from the Harvard Medical School O2 Cluster using R programming (version 4.2.2, DEGreport). Quality control analyses demonstrated that the majority of the samples sequenced by Hybrid Capture had over 20 million total reads. Principal Component Analyses (PCA) and hierarchical clustering analyses of 19,821 protein-coding genes were conducted on the 143 DCIS samples sequenced using Hybrid Capture. PC1 was race (9%). A significant association was observed for follow-up time (PC3, 4.03%). First, we explored the association of the candidate gene ERBB2. The mean transcript abundance for ERBB2 was 11784.37 (case-control p-value=0.08). Preliminary analysis of selected genes of interest did not show statistically significant differences between cases and controls after FDR p-value adjustment. Differential gene expression analyses are ongoing. Conclusion: This study demonstrates the feasibility of generating high-quality RNA-seq data from diverse, archival DCIS samples. Citation Format: Aditi Hazra. Personalized medicine for DCIS [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 6255.

Abstract 7025: Advancing methylation analysis: TAPS v1.0: A fast, scalable, and non-destructive sequencing method

Abstract In the pursuit of advancing multi-cancer early detection, the exploration of cell-free DNA (cfDNA) through methylation profiling has emerged as a promising avenue. However, the realization of its full potential is impeded by limitations inherent in conventional methodologies, notably the 'gold standard' bisulfite conversion methods. These approaches pose challenges such as DNA destruction and the generation of low-complexity sequences resulting from the conversion of unmethylated cytosines to thymine. TET Assisted Pyridine-Borane Sequencing (TAPS) holds the potential as a non-destructive, rapid, and scalable alternative for base-level methylation analysis. In this context, we introduce TET Assisted Pyridine-Borane Sequencing version 1.0 (TAPS v1.0), showcasing advancements to the technology originally introduced by Schuster-Böckler, Song, and their collaborators. TAPS v1.0 transforms 5mC into dihydrouracil (DHU) through a series of oxidation and reduction reactions. Ultimately, DHU is recognized as dU during PCR resulting in C-to-T conversions at 5mC sites, leaving unmethylated cytosines, the majority of cytosines, unchanged. The nondestructive nature of 5mC conversion yields significantly higher library yield and complexity compared to BS-seq. By leveraging Watchmaker's proprietary polymerase evolution platform, a DNA polymerase tolerant to DHU was selected for library amplification post DHU conversion, enabling unimpeded detection of 5mC, particularly in densely packed CpG repeats. Coupled with an optimized End Repair and A-tailing chemistry preceding TAPS v1.0, clinically relevant cfDNA samples of low quality and/or quantity can be analyzed in methylation detection pipelines. To evaluate the performance of TAPS v1.0, human genomic DNA with control spike-ins featuring varying methylation statuses were employed. TAPS v1.0 exhibited significantly higher final library yield and greater complexity compared to BS-seq. Similar outcomes were observed in clinically relevant cfDNA samples. Citation Format: Josh Haimes, Craig Marshall, Travis Sanders, Sam Vogel, Martin Ranik, Eduard Casas, Doug Wendel, Leo Karamanof, Bjarne Faurholme, Abre de Beer, Lee French, Ross Wadsworth, Kristina Giordia, Eric van der Walt, Brian Kudlow. Advancing methylation analysis: TAPS v1.0: A fast, scalable, and non-destructive sequencing method [abstract]. In: Proceedings of the American Association for Cancer Research Annual Meeting 2024; Part 1 (Regular Abstracts); 2024 Apr 5-10; San Diego, CA. Philadelphia (PA): AACR; Cancer Res 2024;84(6_Suppl):Abstract nr 7025.

Developing a Text Messaging Intervention to Prevent Binge and Heavy Drinking in a Military Population: Mixed Methods Development Study.

Alcohol misuse is the fourth leading cause of death in the United States and a significant problem in the US military. Brief alcohol interventions can reduce negative alcohol outcomes in civilian and military populations, but additional scalable interventions are needed to reduce binge and heavy drinking. SMS text messaging interventions could address this need, but to date, no programs exist for military populations. We aimed to develop an SMS text messaging intervention to address binge and heavy drinking among Airmen in Technical Training in the US Air Force. We implemented a 2-phase, mixed methods study to develop the SMS text messaging intervention. In phase 1, a total of 149 respondents provided feedback about the persuasiveness of 49 expert-developed messages, preferences regarding message frequency, timing and days to receive messages, and suggested messages, which were qualitatively coded. In phase 2, a total of 283 respondents provided feedback about the persuasiveness of 77 new messages, including those developed through the refinement of messages from phase 1, which were coded and assessed based on the Behavior Change Technique Taxonomy (BCTT). For both phases, mean persuasiveness scores (range 1-5) were calculated and compared according to age (aged <21 or ≥21 years) and gender. Top-ranking messages from phase 2 were considered for inclusion in the final message library. In phase 1, top-rated message themes were about warnings about adverse outcomes (eg, impaired judgment and financial costs), recommendations to reduce drinking, and invoking values and goals. Through qualitative coding of suggested messages, we identified themes related to warnings about adverse outcomes, recommendations, prioritizing long-term goals, team and belonging, and invoking values and goals. Respondents preferred to receive 1 to 3 messages per week (124/137, 90.5%) and to be sent messages on Friday, Saturday, and Sunday (65/142, 45.8%). In phase 2, mean scores for messages in the final message library ranged from 3.31 (SD 1.29) to 4.21 (SD 0.90). Of the top 5 highest-rated messages, 4 were categorized into 2 behavior change techniques (BCTs): valued self-identity and information about health consequences. The final message library includes 28 BCTT-informed messages across 13 BCTs, with messages having similar scores across genders. More than one-fourth (8/28, 29%) of the final messages were informed by the suggested messages from phase 1. As Airmen aged <21 years face harsher disciplinary action for alcohol consumption, the program is tailored based on the US legal drinking age. This study involved members from the target population throughout 2 formative stages of intervention development to design a BCTT-informed SMS text messaging intervention to reduce binge and heavy drinking, which is now being tested in an efficacy trial. The results will determine the impact of the intervention on binge drinking and alcohol consumption in the US Air Force.

A review on the impact of single-stranded library preparation on plasma cell-free diversity for cancer detection.

In clinical oncology, cell-free DNA (cfDNA) has shown immense potential in its ability to noninvasively detect cancer at various stages and monitor the progression of therapy. Despite the rapid improvements in cfDNA liquid biopsy approaches, achieving the required sensitivity to detect rare tumor-derived cfDNA still remains a challenge. For next-generation sequencing, the perceived presentation of cfDNA is strongly linked to the extraction and library preparation protocols. Conventional double-stranded DNA library preparation (dsDNA-LP) focuses on assessing ~167bp double-stranded mononucleosomal (mncfDNA) and its other oligonucleosomal cell-free DNA counterparts in plasma. However, dsDNA-LP methods fail to include short, single-stranded, or nicked DNA in the final library preparation, biasing the representation of the actual cfDNA populations in plasma. The emergence of single-stranded library preparation (ssDNA-LP) strategies over the past decade has now allowed these other populations of cfDNA to be studied from plasma. With the use of ssDNA-LP, single-stranded, nicked, and ultrashort cfDNA can be comprehensively assessed for its molecular characteristics and clinical potential. In this review, we overview the current literature on applications of ssDNA-LP on plasma cfDNA from a potential cancer liquid biopsy perspective. To this end, we discuss the molecular principles of single-stranded DNA adapter ligation, how library preparation contributes to the understanding of native cfDNA characteristics, and the potential for ssDNA-LP to improve the sensitivity of circulating tumor DNA detection. Additionally, we review the current literature on the newly reported species of plasma ultrashort single-stranded cell-free DNA plasma, which appear biologically distinct from mncfDNA. We conclude with a discussion of future perspectives of ssDNA-LP for liquid biopsy endeavors.

Recovering short DNA fragments from minerals and marine sediments: A comparative study evaluating lysis and isolation approaches

AbstractMarine sediments as excellent climate archives, contain among other biomolecules substantial amounts of extracellular DNA. Through mineral binding, some of the DNA remains protected from degradation which aids its preservation. While this pool of DNA represents genomic ecosystem fingerprints spanning over millions of years, the capability of current DNA extraction methods in recovering mineral‐bound DNA remains poorly understood. We evaluated current sedimentary DNA extraction approaches and their ability to recover short DNA fragments from artificially created DNA‐mineral complexes involving pure clay minerals or quartz, as well as from different types of natural marine sediments. We separately investigated lysis (DNA release) and isolation steps (purification of DNA) comparing five different lysis buffers across two commonly used DNA isolation approaches: silica magnetic beads and liquid‐phase organic extraction and purification. The choice of lysis buffer significantly impacted the amount of recovered mineral‐bound DNA and facilitated selective desorption of DNA fragments. High molarity EDTA and phosphate lysis buffers recovered on average an order of magnitude more DNA from clay minerals than other tested buffers, while both isolation approaches recovered comparable amounts of DNA. In marine sediments, however, liquid‐phase organic extraction caused inhibitory effects in subsequent downstream applications (e.g., PCR), across all assessed DNA extracts, while silica magnetic beads induced inhibition only in half of the tested DNA extracts. Thus, the isolation approach, together with the lysis buffer, played a decisive role in successful library preparation with lysis buffer choice ultimately impacting final library fragment distribution. With this study, we underscore the critical importance of lysis buffer selection to maximize the recovery of mineral‐bound DNA and show its profound impact on recovered fragment lengths in sedimentary DNA extractions, a crucial factor alongside existing isolation approaches in facilitating high‐quality DNA extracts for downstream analysis related to ancient environmental DNA research.

Discovery of Glycation Products: Unraveling the Unknown Glycation Space Using a Mass Spectral Library from In Vitro Model Systems.

The nonenzymatic reaction between amino acids (AAs) and reducing sugars, also known as the Maillard reaction, is the primary source of free glycation products (GPs) in vivo and in vitro. The limited number of MS/MS records for GPs in public libraries hinders the annotation and investigation of nonenzymatic glycation. To address this issue, we present a mass spectral library containing the experimental MS/MS spectra of diverse GPs from model systems. Based on the conceptional reaction processes and structural characteristics of products, we classified GPs into common GPs (CGPs) and modified AAs (MAAs). A workflow for annotating GPs was established based on the structural and fragmentation patterns of each GP type. The final spectral library contains 157 CGPs, 499 MAAs, and 2426 GP spectra with synthetic model system information, retention time, precursor m/z, MS/MS, and annotations. As a proof-of-concept, we demonstrated the use of the library for screening GPs in unidentified spectra of human plasma and urine. The AAs with the C6H10O5 modification, fructosylation from Amadori rearrangement, were the most found GPs. With the help of the model system, we confirmed the existence of C6H10O5-modified Valine in human plasma by matching both retention time, MS1, and MS/MS without reference standards. In summary, our GP library can serve as an online resource to quickly screen possible GPs in an untargeted metabolomics workflow, furthermore with the model system as a practical synthesis method to confirm their identity.

Entrepreneurship skills and marketing library services as correlates of job creation among undergraduates at Tai Solarin University of Education, Ogun State, Nigeria

This study investigated the correlation between entrepreneurship skills, marketing library services, and job creation among undergraduates at Tai Solarin University of Education, Ogun State, Nigeria. Entrepreneurship skills and marketing library services are crucial for job creation among undergraduates; enhancing employability, self-reliance, fostering innovation, and driving economic growth. The study employed descriptive research design, using a self- structured questionnaire for data collection. The population comprised 347 final year library and information science (LIS) undergraduates, with a sample size of 181 with a response rate of 153(84.5%) for data analysis. The findings revealed a highly significant and positive association between entrepreneurship skills and job creation opportunities among the final year undergraduates, with a coefficient (B) for entrepreneurship skills of 0.609 and a standard error of 0.026. Additionally, there is a highly significant and positive relationship between marketing library services and job creation opportunities. Customer service skills and leadership skills had the highest mean values of 3.03 and 3.00, respectively. Respondents indicated strong marketing skills in communication and customer service, with mean scores of 3.63 and 3.44, respectively. Furthermore, the study established strong job opportunities in the fields of librarianship and education and training, each with a mean score of 3.73. The study concluded that entrepreneurship skills and marketing library services are crucial determinants of job creation among final-year LIS undergraduates. It recommended that the university should integrate more extensive entrepreneurship and marketing courses into the LIS curriculum, focusing on practical applications such as customer service and communication skills.

Phage display selection of a Pb(II) specific peptide and its application as a biorecognition unit for colorimetric detection of Pb(II) ions.

Phage display technology employs a library of engineered filamentous M13 viruses infecting only bacteria. In this study, the genuine phage display selection technique was applied to identify a Pb(II) specific peptide. After three rounds of positive selection against Pb(II) coated agarose-based beads and the consecutive negative screenings against interfering metal ions (Al(III), Co(II), Fe(III), Ni(II), and Zn(II)), a final phage library with enhanced Pb(II) binding was obtained. Enzyme Linked Immunosorbent Assay (ELISA) analyses confirmed the selective Pb(II) binding of the enriched viruses. Twenty monoclonal phage plaques were randomly selected, single stranded DNAs (ssDNAs) were isolated and sequenced. Sequencing results revealed four different peptide sequences. Pb9 peptide (KASPYIT) showing the most specific Pb(II) binding was selected for detection studies. Pb9 was synthetically synthesized with additional three cysteine (3xCys) units at C-terminal. Twenty nanometers AuNPs were functionalized with Pb9-3xCys peptides through Au-thiol (Au-S) interaction. A colorimetric Pb(II) detection system was validated using the engineered peptide-AuNP complex at a calculated LOD of around 11nM (3σ/k, n=6) for the case study. The detection system was Pb(II) selective over various metal ions (Ag(II), Al(III), Au(III), Cd(II), Co(II), Cr (III), Cu(II), Fe(III), Hg(II), Mg(II), Mn(II), Ni(II), and Zn(II)). Such metal ion specific peptides can be further studied to develop simple, user friendly and cost-effective tools to design alternative detection and bioremediation systems for a circular economy.

A group B Streptococcus indexed transposon mutant library to accelerate genetic research on an important perinatal pathogen.

Group B Streptococcus (GBS) is a significant global cause of serious infections, most of which affect pregnant women, newborns, and infants. Studying GBS genetic mutant strains is a valuable approach for learning more about how these infections are caused and is a key step toward developing more effective preventative and treatment strategies. In this resource report, we describe a newly created library of defined GBS genetic mutants, containing over 1,900 genetic variants, each with a unique disruption to its chromosome. An indexed library of this scale is unprecedented in the GBS field; it includes strains with mutations in hundreds of genes whose potential functions in human disease remain unknown. We have made this resource freely available to the broader research community through deposition in a publicly funded bacterial maintenance and distribution repository.

Evaluation of the Thermo Fisher Oncomine Myeloid MRD DNA Assay on the Ion Torrent Platform

Introduction Malignant myeloid diseases are a group of blood cancers that arise from genetic changes in hematopoietic cells in the bone marrow or peripheral blood. The presence of measurable residual disease (MRD), or detection of abnormal cells in the body following completion of therapy, is associated with a higher risk of disease relapse, and decreased overall survival (Schuurhuis 2018). Next-generation sequencing (NGS) can be leveraged as a high-sensitivity, minimally invasive method for detection of MRD. The Ion Torrent Oncomine Myeloid MRD Assay (Myeloid MRD) is a targeted NGS assay designed for the detection of single nucleotide variants (SNVs) and insertions/deletions in a curated selection of clinically relevant myeloid disease-related genes at low limits of detection. Assay performance was evaluated using normal and diseased clinical blood and bone marrow samples and NGS reference materials. Methods The Myeloid MRD DNA panel targets hotspot regions across 33 genes and uses Ion AmpliSeq HD technology with unique molecular tagging to suppress sequencing artifacts and improve analytical sensitivity. Briefly, DNA was extracted from whole blood or bone marrow samples and 120 ng of nucleic acid was used as input for the assay. The DNA was divided between 2 polymerase chain reactions (PCR), where it served as the template for target amplifications, before combining reaction products by sample. Library preparation followed the standard Ion AmpliSeq HD protocol, where amplicons were partially enzymatically digested, and a second round of amplification was performed using barcoded primers. Following bead purification, the final library was measured by quantitative PCR (qPCR) to inform final library loading onto the sequencer. Six libraries were pooled and sequenced on an Ion 550 chip using the Ion GeneStudio S5 System. Primary and secondary analyses were performed using the Torrent Suite software, and results were exported to the Ion Reporter software for variant calling. From the variant calling results, a probit model was utilized to determine the variant allele frequency (VAF) at which the assay achieved 90% power to call an expected variant, defining the limit of detection (LoD). Other parameters evaluated included analytical sensitivity, analytical specificity, positive predictive value (PPV), accuracy, and precision. Results Overall, consistent, high quality on-target reads were observed across 22 sequencing runs with a median Ion Sphere Particle (ISP) loading of 90%, usable sequence of 59%, and over 130 million total reads per chip. When evaluating individual samples according to the vendor-recommended number of mapped reads (&amp;gt; 10 million), 98.3% of samples tested passed quality control (QC). Additional sample QC metrics established during assay validation included thresholds of &amp;gt; 80% reads on-target, &amp;gt; 80% uniformity of base coverage, and a mean read length of at least 80 base pairs. The estimated LoD for the assay was 0.1% VAF for small variants (SNVs, insertions/deletions).Above the LoD, the assay demonstrated 96.03% sensitivity, 99.89% accuracy of base-calling, 100% specificity, and 98.83% PPV.Additionally,a comparison was performed usingmatched whole blood and bone marrow samples collected from patients with acute myeloid leukemia (AML) to evaluate the concordance of variant calling. It was determined that the same variants were identified across both sample types, although at higher allele frequency in bone marrow compared to whole blood. Conclusion This evaluation of the Myeloid MRD assay demonstrates that it is a precise and accurate assay with sensitivity that is acceptable for MRD analysis. Notably, variant detection was concordant between whole blood and bone marrow sample types collected from AML patients, with bone marrow providing improved sensitivity for low-frequency variant detection. The user-friendly workflow and integrated informatics pipeline offer opportunities for streamlined testing and interpretation of results. The Ion Torrent Myeloid MRD DNA assay is a powerful new tool for MRD detection across a growing myeloid disease target landscape.

Local food procurement by hospitals: a scoping review.

Local food procurement by hospitals is gaining traction as governments and advocacy groups seek to influence food systems and strengthen local communities, but there is little empirical evidence as to its practical application or efficacy. The aims of this review were to describe the extent, range and nature of local food procurement models in healthcare foodservices, and to understand the barriers and enablers to implementation, including from the perspective of stakeholders across the supply chain. A scoping review was conducted following the protocol published in the Open Science Framework Registration (DOI: 10.17605/OSF.IO/T3AX2). Five electronic databases were searched for the following concepts: 'hospital foodservice', 'local food procurement practices', 'the extent, range and nature' and/or 'the barriers and enablers of procurement'. Eligible peer-reviewed original research published in English from the year 2000 was included following a two-step selection process. The final library included nine studies. Most studies (7 of 9) were from the United States. Three studies used survey methods and reported high rates (58%-91%) of US hospital participation in local food procurement. Studies offered minimal description of local procurement models, but two models, conventional ('on-contract') or off-contract, were typically used. Barriers to local food procurement included restricted access to suitable local food supply, limited kitchen resources and inadequate technology to trace local food purchase thereby limiting evaluation capabilities. Enablers included organisational support, passionate champions and opportunistic, incremental change. There is a paucity of peer-reviewed studies describing local food procurement by hospitals. Details of local food procurement models were generally lacking: categorisable as either purchases made 'on-contract' via conventional means or 'off-contract'. If hospital foodservices are to increase their local food procurement, they require access to a suitable, reliable and traceable supply, that acknowledges their complexity and budgetary constraints.

SIGnature Libraries: A roadmap for the formation of special interest group libraries.

"SIGnature Libraries" channel the dynamism of academic society-based special interest groups (SIG) to systematically identify and provide user-oriented access to essential literature for a subspecialty field in a manner that keeps pace with the field's continuing evolution. The libraries include literature beyond clinical trial data to encompass historical context, diagnostic conceptualization, and community organization materials to foster a holistic understanding of how neurologic conditions affect individuals, their community, and their lived experience. Utilizing a modified-Delphi approach, Child Neurology Society's Cerebral Palsy (CP) SIG (n = 75) administered two rounds of literature submissions and ratings. A final review by an 11-member international advisory group determined threshold ratings for resource inclusion and the library's final structure. Seventy-nine articles were submitted for the first Delphi round and 22 articles for the second Delphi round. Survey response rates among SIG members were 29/75 for the first round and 24/75 for the second round. The advisory board added additional articles in the final review process in view of the overall project goal. A total of 60 articles were included in the final library, and articles were divided into seven sections and stratified by rating scores. A process for ongoing revisions of the library was determined. The library will be published on the Child Neurology Society website and made publicly accessible. The CP SIGnature Library offers learners an unprecedented resource that provides equitable access to latest consensus guidelines, existing seminal datasets, up-to-date review articles, and other patient care tools. A distinctive feature of the library is its intentional large scope and depth, presented in a stratified fashion relative to the consensus-determined importance of each article. Learners can efficiently navigate the library based on individual interests and goals, and the library can be used as core curriculum for CP education.

Blocking Abundant RNA Transcripts by High-Affinity Oligonucleotides during Transcriptome Library Preparation

BackgroundRNA sequencing has become the gold standard for transcriptome analysis but has an inherent limitation of challenging quantification of low-abundant transcripts. In contrast to microarray technology, RNA sequencing reads are proportionally divided in function of transcript abundance. Therefore, low-abundant RNAs compete against highly abundant - and sometimes non-informative - RNA species.ResultsWe developed an easy-to-use strategy based on high-affinity RNA-binding oligonucleotides to block reverse transcription and PCR amplification of specific RNA transcripts, thereby substantially reducing their abundance in the final sequencing library. To demonstrate the broad application potential of our method, we applied it to different transcripts and library preparation strategies, including YRNAs in small RNA sequencing of human blood plasma, mitochondrial rRNAs in both 3′ end sequencing and long-read sequencing, and MALAT1 in single-cell 3′ end sequencing. We demonstrate that the blocking strategy is highly efficient, reproducible, specific, and generally results in better transcriptome coverage and complexity.ConclusionOur method does not require modifications of the library preparation procedure apart from simply adding blocking oligonucleotides to the RT reaction and can thus be easily integrated into virtually any RNA sequencing library preparation protocol.