Tag: MOI

In late May I had the honor of speaking with Sucharit Bhakdi, MD regarding dsDNA contamination in the vaccines. First, we need to do some numbers. The 100ug Moderna shots deliver over 40 trillion mRNA molecules (14T for Pfizer) and the number of LNPs per shot is reported to be…

Sample collection and cDNA synthesis In this study, we used porcine alveolar macrophages (PAMs) that were isolated from 16 different animals between the years 2011 and 2022, and subsequently stored. The PAMs were isolated by bronchoalveolar lavage (BAL) from lungs of euthanized 3-week-old healthy conventional piglets from a PRRSV negative…

Assembly of T4 artificial viral vectors T4-AVVs were assembled by sequential incorporation of purified biomaterials to generate a virus structural mimic (Fig. 2a and Supplementary Movie 1). Starting with an empty capsid shell purified from E. coli infected by the neck-minus and tail-minus T4 phage mutant (10-amber.13-amber.HocΔ.SocΔ T4) (Supplementary Fig. 1a), a pentameric…

Recombinant ASFVs with mosaic genomes of genotype I and II viruses detected from field samples in China During our surveillance in China, we isolated three ASFVs from pig samples collected in Jiangsu province, Henan province, and Inner Mongolia Autonomous Region, respectively, and designated them Pig/Jiangsu/LG/2021 (JS/LG/21), Pig/Henan/123014/2022 (HeN/123014/22), and Pig/Inner…

Screening strategy for identifying viral inhibitors of the cGAS/STING pathway Vaccinia virus is a large cytoplasmic DNA virus with a 190 kilobase pairs (kbp) genome that encodes over 200 proteins21,22. MVA has an approximately 30-kbp deletion from its parental vaccinia genome, resulting in the loss of many immune-modulatory viral genes15….

RNA-seq analyses of adult and weanling brain microvessels identify candidate gene regulators of LACV infection We first sought to identify candidate gene regulators that contribute to age-dependent susceptibility to LACV. Weanling (W) and adult (A) mice were treated with either poly I:C (PI) for 3 h (as a surrogate for a…

Identification of MDV presence using open access sequencing data Metagenomic next-generation sequencing (NGS) data for field mosquitoes in China were searched from the NCBI SRA database (www.ncbi.nlm.nih.gov/sra) and CNGB Sequence Archive database (db.cngb.org/cnsa/) using the keywords “mosquito OR Anopheles OR Aedes OR Culex”. Followed by removing the datasets with less…

Plasmid design The information on each plasmid for combinatorial libraries is indicated in Supplementary Table S1. Plasmids used in this study to generate combinatorial libraries for SpCas9, enAsCas12a, CHyMErA, enAsCas12a (dual-gRNA), CHyMErA.v2, multiSPAS were deposited to Addgene (IDs: 189632, 189633, 189634, 189635, 189636, 189637, respectively). 3Cs oligonucleotide design The protocol…

Ethics statement All experiments in this study comply with guidelines of the U.S. Public Health Service and were approved by the Institutional Biosafety Committee at the University of Pittsburgh. All animal husbandry and experimental procedures were conducted in accordance with U.S. Public Health Service policy and approved by the Institutional…

Ethics declaration Animal study was performed following the Guide for the Care and Use of Laboratory Animals. Animal experiment was approved by the Institutional Animal Care and Use Committee of Colorado State University (protocol number 1090) in advance and conducted in compliance with the Association for the Assessment and Accreditation…

CLIP enables high-efficiency knock-in into an essential locus To create CLIP, we leverage integrase-deficient lentiviruses (IDLV) to deliver an HDR donor template as a viral RNA genome to cells25,26. IDLV consists of lentiviral components but with a D64V mutation in the integrase, maintaining this enzyme’s ability to package and nuclear…

All methods were performed in accordance with institutional guidelines and regulations. CRISPR-Cas9 protein and guide RNA reagents All Cas9 proteins used in this study were derived from the prototypic Streptococcus pyogenes species. Cas9 protein was produced as described in Seamon et al.33 and contained a N-terminal nuclear localization signal and…

Cell lines and tissue culture The murine 17-CL1 cell line (derived from 3T3 cells, BEI Resources, Cat. No. NR-53719) was maintained in Dulbecco Modified Eagle Medium (DMEM, Life Technologies) supplemented with 10% Fetal Bovine Serum (FBS, Gemini Bio-Products), L-glutamine (Invitrogen), and Sodium pyruvate (Invitrogen). The NCTC clone 1469 cell line…

Ethics Human blood was obtained from healthy donors, approved by the Regional Ethics Committee and Imperial College Healthcare NHS Trust Tissue Bank (Regional Ethics Committee approval no. 17/WA/0161, Imperial College Healthcare Tissue Bank Human Tissue Authority license no. 12275, and Imperial College Research Ethics Committee no. 19IC5166). All samples were…

Cell culture and reagents HEK293 and HeLa cells (American Type Culture Collection) were cultured in Dulbecco’s modified Eagle’s medium (DMEM) supplemented with 10% fetal calf serum, 2 mM GlutaMAX‐1, 100 μg/mL streptomycin and 100 U/mL penicillin (complete growth medium) at 37 °C under 5% CO2. HeLa cells stably expressing GFP-TIFA were previously described8. Cells…

Purification of T7Select 10-3b phages The T7Select 10-3b phage vector (Novagen, Merck) was used in this study. Non-recombinant/recombinant T7Select 10-3b phages were purified from the lysate of infected E. coli strain BL5403 by PEG-8000 precipitation as per the method described in T7 select system manual. The pellet was suspended in…

Poxvirus vectors for directed evolution of GPCRs in mammalian cells Monoclonal and homogenous expression of the gene of interest are critical prerequisites for selections in directed evolution that are difficult to achieve in mammalian cells with common expression strategies such as transient transfection. To reach both equal gene dosage and…

Motivation for developing a sparse RDB Computational approaches for discovering the etiologies of rare diseases typically depend on the analysis of a heterogeneous set of files, each of which can be very large and follow a distinct convention. Genotypes, for example, are ordinarily stored in VCFs containing data for one…

With an easy-to-use instrument to encapsulate single cells in picolitre-volume water droplets (picodroplets) at high throughput rates of thousands of droplets per second, picodroplet microfluidics is quickly emerging as the technology of choice for single-cell analysis. This highly effective and delicate method has important benefits in terms of cost and…

Screening for hyper-potent shRNA against SARS-CoV-2 We parsed the SARS-CoV-2 genome into a series of potential short hairpin RNA (shRNA) targets (Supplemental Fig. 1). This process was conducted by tiling the genome with overlapping 50 nucleotide-long sequences, each shifted by a single nucleotide from the other. The region targeted by each…

Genetic screen identifies modulators of Tun-mediated neuronal death To identify genes that ameliorate neuronal cell death caused by Tun-induced ER stress, we performed a loss-of-function pooled genetic screen using our druggable genome sgRNAs library. We first generated a clonal iPSC line that expresses Cas9 under the GAPDH promoter (Fig. 1A). We…

Cell lines Immortalised HA1E (hTERT and SV40ER) and tumourigenic HA1ER cells (hTERT, SV40ER and HRAS-G12V) from stepwise tumourigenesis models generated from normal human embryonic kidney cells were obtained from Dr. Hahn (Broad Institute of MIT and Harvard, Cambridge, USA). Cells were cultured in DMEM (1 g/L glucose) (Gibco, cat. #10567014) supplemented…

Phylogenetic inference To analyze the phylogenetic relationship of the TBEV sequences obtained from ticks in the Netherlands (GenBank accession numbers LC171402 and MH021184)21,22 with known TBEV-Eu strains, all TBEV-Eu sequences encompassing the complete coding sequence were selected from GenBank (n = 88, retrieved on September 18, 2021). Multiplicities, synthetic and clonal sequences…

This research complies with all relevant ethical regulations approved by the Human Investigation Review Board of Albert Szent-Györgyi Clinical Centre of the University of Szeged and the National Biodiversity Authority (NBA) of India. Permission for the faecal sample collection was obtained from the Human Investigation Review Board of Albert Szent-Györgyi…

doi: 10.1002/mco2.208. eCollection 2023 Feb. Zongzhi Liu  1   2   3 , Xiang Gao  4 , Chuanwen Kan  2   5 , Lingyu Li  1   2   3 , Yuan Zhang  6 , Yibo Gao  7 , Shengyuan Zhang  4 , Liangji Zhou  6 , Hui Zhao  6   8   9 , Mingkun Li  2  …

Introduction Lymphoma, a cancer characterized by a malignant tumor of the immune system that originates in the lymph nodes or lymphoid tissue, is one of the most common cancers worldwide. According to GLOBOCAN, the incidence of non-Hodgkin lymphoma (NHL) in both males and females ranked top ten among all cancers…

Viral strains utilized in this work The following viral strains have been used in this study: Pandoravirus neocaledonia3, Pandoravirus macleodensis3, Pandoravirus kuranda (this study, ON887157), Mollivirus kamchatka26, Pithovirus sibericum27 and Mimivirus reunion28. Cloning of DNA constructs All primers used in this study are listed in Table S1. All vectors used in…

CDKs support SARS-CoV-2 RdRp-mediated gene expression To identify cellular proteins that modulate the SARS-CoV-2 RNA synthesis, we first performed immunoprecipitation coupled with mass spectrometry (MS) to determine cellular proteins that associate with SARS-CoV-2 RdRp. Briefly, Flag-tagged nsp7, Flag-tagged nsp8, and Flag-tagged nsp12 plasmids were co-transfected into HEK293T cells, and the…

Cells and reagents HEK293T and U87 cells were maintained in DMEM medium (Gibco) supplemented with 10% fetal bovine serum (FBS, Labtech) and 100 U ml−1 penicillin plus 100 μg ml−1 streptomycin (Pen/Strep; Gibco). THP-1-IFIT1 cells that had been modified to express Gaussia luciferase under the control of the IFIT1 promoter were described previously62. THP-1…

Lai, L. et al. Production of alpha-1,3-galactosyltransferase knockout pigs by nuclear transfer cloning. Science 295, 1089–1092. doi.org/10.1126/science.1068228 (2002). ADS  CAS  Article  PubMed  Google Scholar  Kuwaki, K. et al. Heart transplantation in baboons using alpha1,3-galactosyltransferase gene-knockout pigs as donors: initial experience. Nat. Med. 11, 29–31. doi.org/10.1038/nm1171 (2005). CAS  Article  PubMed  Google…

Coronavirus disease 2019 (COVID-19) vaccines have played a critical role in reducing transmission of the severe acute respiratory syndrome coronavirus 2 (SARS-CoV-2) transmission. However, with emerging reports of waning vaccine efficacy, there remains an urgent need to develop prophylactic measures against COVID-19. In a recent study published on the bioRxiv*…

Virology: Module 2 Flashcards | Quizlet   attachment/entry, translation of viral mRNA into viral proteins, replication, assembly, release of viruses from cell steps of infection for viruses? a cell possesses the functional receptor for a virus – virus can bind, possibly enter, but replication isn’t guaranteed a cell does not…

INTRODUCTION Dengue is the most common mosquito-borne viral disease globally (1). This acute disease, which can be life-threatening, is caused by four different dengue viruses (DENVs) (DENV-1, DENV-2, DENV-3, and DENV-4). An estimated 390 million people are infected with these DENVs annually (2), and populations throughout the tropics face frequent…

Significance Interferon (IFN)-mediated antiviral responses serve as the first line of the host innate immune defense against viral infection. Here we identify a previously uncharacterized protein designated phospholipase A2 inhibitor and LY6/PLAUR domain-containing protein (PINLYP), which is essential for embryonic development and plays an important role in type I IFN…

Health experts in Kenya aim to reduce the number of children born with sickle cell disease — an inherited blood disorder — by more than 12 times through public education and awareness, as the country premiered a guideline for controlling and managing the disease. The guideline titled Sickle Cell Disease…