Tag: NEB

Debian — Details of package cp2k in bookworm

Ab Initio Molecular Dynamics CP2K is a program to perform simulations of solid state, liquid, molecular and biological systems. It is especially aimed at massively parallel and linear scaling electronic structure methods and state-of-the-art ab-initio molecular dynamics (AIMD) simulations. CP2K is optimized for the mixed Gaussian and Plane-Waves (GPW) method…

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Detection of DNA methylation signatures through the lens of genomic imprinting

Animals and samples The study included 10 pigs, 8 pigs were bred at the INRAE experimental farm (doi.org/doi.org/10.15454/1.5572415481185847E12) and 2 pigs come from breeding organizations in accordance with the French and European legislation on animal welfare. The animals belong to the same family, except for one LW animal. Animals were…

Continue Reading Detection of DNA methylation signatures through the lens of genomic imprinting

Seroprevalence and Molecular Characterization of B. abortus

Introduction Brucellosis is a zoonotic disease caused by Brucella spp., a gram-negative facultative intracellular coccobacillus.1 These microorganisms can infect livestock, wildlife, and humans, causing significant public concern and substantial agricultural economic loss. Currently, at least six novel Brucella species have been identified: Brucella pinnipedialis, Brucella ceti,2 Brucella papionis,3 Brucella microti,4…

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Frontiers Publishing Partnerships | Construction of recombinant adenovirus-5 vector to prevent replication-competent adenovirus occurrence

Introduction In recent years, recombinant adenoviral vectors have been used in different fields of biomedical sciences such as in vitro and in vivo gene transfer, vaccine development, and gene therapy (Russell, 2000; Mitani and Kubo, 2002). Multiple features of recombinant adenoviral vectors such as high packaging capacity for transgene insertion,…

Continue Reading Frontiers Publishing Partnerships | Construction of recombinant adenovirus-5 vector to prevent replication-competent adenovirus occurrence

Genomic hypomethylation in cell-free DNA predicts responses to checkpoint blockade in lung and breast cancer

Lung cancer ICB cohort Advanced non-small cell lung carcinoma patients who were treated with anti-PD-1/PD-L1 monotherapy at Samsung Medical Center, Seoul, Republic of Korea were enrolled for this study. The present study has been reviewed and approved by the Institutional Review Board (IRB) of the Samsung Medical Center (IRB no….

Continue Reading Genomic hypomethylation in cell-free DNA predicts responses to checkpoint blockade in lung and breast cancer

Evidence of an intracellular creatine-sensing mechanism that modulates creatine biosynthesis via AGAT expression in human HAP1 cells

Cell lines and reagents HAP1 cells used to generate the reporter cells were grown from a frozen stock of HAP1 (C859) passage 6 purchased from Horizon Discovery LTD (Cambridge, UK). Iscove’s media (Ref: 098–150, Wisent) supplemented with 10% Fetal Bovine serum (Multicell) was used to grow HAP1 cells. All cells…

Continue Reading Evidence of an intracellular creatine-sensing mechanism that modulates creatine biosynthesis via AGAT expression in human HAP1 cells

A high-resolution transcriptomic and spatial atlas of cell types in the whole mouse brain

Mouse breeding and husbandry All experimental procedures related to the use of mice were approved by the Institutional Animal Care and Use Committee of the AIBS, in accordance with NIH guidelines. Mice were housed in a room with temperature (21–22 °C) and humidity (40–51%) control within the vivarium of the AIBS…

Continue Reading A high-resolution transcriptomic and spatial atlas of cell types in the whole mouse brain

Benchmarking DNA isolation methods for marine metagenomics

Scheme of the study The overall pipeline of our study is presented in Fig. 1 and is described in detail in the methods section. We processed three types of samples: fresh water, sea sediment, and digestive system of a marine invertebrate M. gigas (“gut flora”). These samples were treated in triplicates…

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Biosensors | Free Full-Text | CRISPR/Cas12a-Based Detection Platform for Early and Rapid Diagnosis of Scrub Typhus

1. Introduction Orientia tsutsugamushi (OT) is an obligate intracellular parasite bacteria and the causative agent of scrub typhus (ST), which is associated with acute febrile illness (AFI) [1] and transmitted by mites through an infected chigger bite (in the larval stage). This disease, which was earlier believed to be endemic…

Continue Reading Biosensors | Free Full-Text | CRISPR/Cas12a-Based Detection Platform for Early and Rapid Diagnosis of Scrub Typhus

Employment of a high throughput functional assay to define the critical factors that influence vaccine induced cross-variant neutralizing antibodies for SARS-CoV-2

Study design, ethical statement, and sample collection The vaccinated participants were recruited under the COVID-19 PROTECT study (2012/00917). The convalescent plasma samples were collected from subjects who were diagnosed with COVID-19 by positive polymerase chain reaction (PCR) results (study 2020/00120). All participants provided written informed consent in accordance with the…

Continue Reading Employment of a high throughput functional assay to define the critical factors that influence vaccine induced cross-variant neutralizing antibodies for SARS-CoV-2

Resin acids play key roles in shaping microbial communities during degradation of spruce bark

Bark preparation Spruce bark was obtained from the Iggesund pulp and paper mill (Iggesund, Holmen AB, Sweden), from a bark pile resulting from stripping of spruce logs at the mill after harvest, with the average age of trees at harvest being ~70 years. The bark was left to dry at…

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Genome sequence and characterization of a novel Pseudomonas putida phage, MiCath

Bacterial strains We used P. putida strains S12, DOT-T1E, F1 (kindly gifted by Grant Rybnicky), ATCC 12633 (purchased from ATCC), JUb85 (kindly provided by Samuel Buck), EM383 (kindly gifted by Huseyin Tas), p106 (kindly provided by Carey-Ann Burnham), and KT2440 (obtained from lab stocks). An overnight culture of each P….

Continue Reading Genome sequence and characterization of a novel Pseudomonas putida phage, MiCath

Generating high-quality plant and fish reference genomes from field-collected specimens by optimizing preservation

Sample collection A total of nine species of marine fish were collected across three different sampling days (September 7th, 9th, and 12th 2022) under IACUC Animal Use Protocol S12219 (Supplementary Data 1). Six species were collected using a speargun donated by a local fisher. Fish were transported back to shore, euthanized,…

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DNA polymerases in precise and predictable CRISPR/Cas9-mediated chromosomal rearrangements | BMC Biology

Cell culture The human endometrial carcinoma HEC-1-B cells were cultured in the modified Eagle’s medium (MEM) supplemented with 10% fetal bovine serum (FBS) and 1% penicillin-streptomycin at 37°C in a 5% (v/v) CO2 incubator. The human embryonic kidney HEK293T cells were cultured in the Dulbecco’s modified Eagle’s medium (DMEM) supplemented…

Continue Reading DNA polymerases in precise and predictable CRISPR/Cas9-mediated chromosomal rearrangements | BMC Biology

Genomic DNA extraction optimization and validation for genome sequencing using the marine gastropod Kellet’s whelk [PeerJ]

Introduction Rapidly advancing next generation sequencing technologies such as whole genome sequencing, genotyping-in-thousands by sequencing (GT-seq), and restriction-site-associated DNA sequencing (RAD-seq), are becoming more available and affordable for non-model organisms (Park & Kim, 2016; Ellegren, 2014; Van Wyngaarden et al., 2017; Bootsma et al., 2020). However, applying these technologies to…

Continue Reading Genomic DNA extraction optimization and validation for genome sequencing using the marine gastropod Kellet’s whelk [PeerJ]

Prime editing-mediated correction of the CFTR W1282X mutation in iPSCs and derived airway epithelial cells

Abstract A major unmet need in the cystic fibrosis (CF) therapeutic landscape is the lack of effective treatments for nonsense CFTR mutations, which affect approximately 10% of CF patients. Correction of nonsense CFTR mutations via genomic editing represents a promising therapeutic approach. In this study, we tested whether prime editing,…

Continue Reading Prime editing-mediated correction of the CFTR W1282X mutation in iPSCs and derived airway epithelial cells

Common analysis of direct RNA sequencinG CUrrently leads to misidentification of m5C at GCU motifs

Introduction Oxford Nanopore Technologies (ONT) direct RNA sequencing (Fig 1A) enables detection of RNA modifications. A modified base produces an altered electrical current and/or dwell time relative to a canonical base that can be detected with algorithms (Garalde et al, 2018; Smith et al, 2019; Workman et al, 2019). Figure…

Continue Reading Common analysis of direct RNA sequencinG CUrrently leads to misidentification of m5C at GCU motifs

sgRNA Synthesis Reagents and Kits Products

Whether you are performing your first cloning experiment or constructing multi-fragment DNA assemblies, NEB® has the solution for you. Our high quality reagents are available for every workflow, including popular DNA assembly methods such as NEBuilder® HiFi DNA Assembly and NEBridge® Golden Gate Assembly. We also offer solutions for automation, site-directed…

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What is the Difference Between pBR322 and pUC19

The main difference between pBR322 and pUC19 is that pBR322 has two origins of replication, one derived from the ColE1 plasmid and another from the pMB1 plasmid, whereas pUC19 has a single origin of replication derived from the pMB1 plasmid. pBR322 and pUC19 are both important plasmids in molecular biology….

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Chromatin priming elements direct tissue-specific gene activity before hematopoietic specification

Introduction The development of multicellular organisms requires the activation of different gene batteries which specify the identity of each individual cell type. Such shifts in cellular identity are driven by shifts in the gene regulatory network (GRN) consisting of transcription factors (TFs) binding to the enhancers and promoters of their…

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A single pseudouridine on rRNA regulates ribosome structure and function in the mammalian parasite Trypanosoma brucei

Cell growth and transfections Procyclic form (PCF) T. brucei, strain 29-1354, which carries integrated genes for the T7 polymerase and the tetracycline repressor, was grown in SDM-79 medium supplemented with 10% fetal calf serum, in the presence of 50 μg/ml hygromycin. Cells were grown in the presence of 15 μg/ml G418 for…

Continue Reading A single pseudouridine on rRNA regulates ribosome structure and function in the mammalian parasite Trypanosoma brucei

GDF11 slows excitatory neuronal senescence and brain ageing by repressing p21

Experimental model and subject details Mice Male ICR mice (Laboratory Animal Center of Zhejiang Academy of Medical Sciences) at age of 3 months (M), 9 M and 36 M, male C57BL/B6 wild‐type (WT) mice (Shanghai Slac Laboratory) at age of 3 M and 10 M, male GDF11-flox mice (GDF11f/f, mice carrying the “floxed” GDF11…

Continue Reading GDF11 slows excitatory neuronal senescence and brain ageing by repressing p21

DRIP-qPCR validation – Mendeley Data

Description 1. HEK293 cells were obtained from the National Collection of Authenticated Cell Culture of China (accession number: SCSP-5014) and cultured in DMEM supplemented with 10% FBS and penicillin/streptomycin (100U/ml) at 37°C with 5%CO2. Expression vector carrying NOTCH2NLC exon1 with different numbers of GGC repeats were prepared and transfected into…

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DNA and RNA Kits for Faster, Easier NGS Library Prep

IPSWICH, MA — New England Biolabs (NEB®) announced the release of the NEBNext UltraExpress DNA and RNA Library Prep Kits for next generation sequencing on the Illumina platform. These new kits are designed to generate high-quality, high-yield libraries with a faster, easier workflow, enabling RNA library prep in a single…

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Deploy CP2K on an Azure virtual machine

This article describes the steps for running a CP2K application on a virtual machine (VM) deployed on Azure. It also presents the performance results of running CP2K on single-node and multi-node VM configurations. CP2K is a quantum chemistry and solid-state physics software package that can perform atomistic simulations of solid…

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A Cre-dependent massively parallel reporter assay allows for cell-type specific assessment of the functional effects of non-coding elements in vivo

Animal models All procedures involving animals were approved by the Institutional Animal Care and Use Committee (IACUC) at Washington University in St. Louis, MO. Veterinary care and housing was provided by the veterinarians and veterinary technicians of Washington University School of Medicine under Dougherty lab’s approved IACUC protocol. All protocols…

Continue Reading A Cre-dependent massively parallel reporter assay allows for cell-type specific assessment of the functional effects of non-coding elements in vivo

RNAseq from P4 murine whole kidneys with Polycystic Kidney Disease

Snap frozen Postnatal day4 murine kidney tissues were crushed using micropestles in RLT Buffer and RNA was extracted from these tissues using QiaShredders followed by Qiagen RNAeasy Mini Kits according to manufacturer’s protocol. RNA concentration and integrity were determined by Aligent bioanalyzer. cDNA library preparation was carried out using NEB…

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Establishment of RT-RPA-Cas12a assay for rapid and sensitive detection of human rhinovirus B | BMC Microbiology

Clinical samples Thirty nasopharyngeal aspirates were collected from individuals with symptoms of respiratory tract infections, and tested positive for HRV-A (5 clinical samples), HRV-B (20 clinical samples), or HRV-C-positive (5 clinical samples) based on the method, as previously described [27]. Ten other specimens were tested negative for HRV-B, including human…

Continue Reading Establishment of RT-RPA-Cas12a assay for rapid and sensitive detection of human rhinovirus B | BMC Microbiology

IJMS | Free Full-Text | CRISPR/Cas9 Directed Reprogramming of iPSC for Accelerated Motor Neuron Differentiation Leads to Dysregulation of Neuronal Fate Patterning and Function

1. Introduction As the world population ages, neurodegenerative diseases are increasing. The outcome of these diagnoses varies but can lead to fatality 50% of the time within 15–20 months [1]. Treatments for neurodegeneration, in particular motor neuron (MN) diseases, are restricted due to the limitations of motor neuron repair and…

Continue Reading IJMS | Free Full-Text | CRISPR/Cas9 Directed Reprogramming of iPSC for Accelerated Motor Neuron Differentiation Leads to Dysregulation of Neuronal Fate Patterning and Function

FAT10 is phosphorylated by IKK{beta} to inhibit the antiviral type-I interferon response

Introduction The innate immune system represents the host´s first-line defense against viral infections (Koyama et al, 2008). In this context, IFN-I are produced by all nucleated cells and represent the principal cytokines that counteract viral replication (Ivashkiv & Donlin, 2014). In fact, several receptors of the infected cell can recognize…

Continue Reading FAT10 is phosphorylated by IKK{beta} to inhibit the antiviral type-I interferon response

Mapping Mammalian 3D Genome Interactions with Micro-C-XL

A protocol for mapping the three-dimensional genome organization with nucleosome resolution using the genome-wide chromosome conformation capture method Micro-C-XL is presented here. This protocol is significant as it shows high resolution chromosome loops and other short-range interaction features. The main advantage of this technique is the mapping of the 3D…

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Expression profiles of circRNAs and biomarkers for COPD

Introduction Chronic obstructive pulmonary disease (COPD) is a chronic inflammatory lung disease featured by airflow obstruction with high prevalence, morbidity, and mortality. It is reported that COPD is the third leading cause of death globally at present, and the incidence of COPD in adults over 40 years is more than…

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Enduring Basics and Evolving Enzyme Kits

By MaryAnn Labant Next-generation sequencing (NGS) is increasingly employed to dive deeper into complex biological mechanisms in a myriad of disciplines. But as with most methodologies, the quality of the input material determines the quality of the resultant data. High-quality sample preparation is the foundation of NGS, and the choices…

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The DNA-binding induced (de)AMPylation activity of a Coxiella burnetii Fic enzyme targets Histone H3

Statistics and reproducibility Anti-AMP IP for LC-MS/MS analysis was performed in three independent biological replicates (n = 3). TSA assay data represents technical triplicates. Time-resolved (de)AMPylation analyzed by LC-MS was performed as biological triplicates. CD measurements were performed in technical triplicates. Anisotropy data is shown as technical triplicates. Anisotropy measurements were repeated…

Continue Reading The DNA-binding induced (de)AMPylation activity of a Coxiella burnetii Fic enzyme targets Histone H3

DNA hypomethylation characterizes genes encoding tissue-dominant functional proteins in liver and skeletal muscle

Overview of this study In this study, we measured the DNA methylome from mouse liver and skeletal muscle, integrated the data with the transcriptome and proteome of these mouse tissues22,23, and examined how tissue-dominant protein and gene expression were associated with DNA hypomethylation (Fig. 1). In this study, we measured DNA…

Continue Reading DNA hypomethylation characterizes genes encoding tissue-dominant functional proteins in liver and skeletal muscle

Comparative analysis of shotgun metagenomics and 16S rDNA sequencing of gut microbiota in migratory seagulls [PeerJ]

Introduction The microbiomes of different host organisms are currently described by using culture-independent sequencing methods (Harvey & Holmes, 2022). Shotgun metagenomic and 16S rDNA sequencing methods are commonly used to identify the taxonomic composition of microbial communities in several studies (de Melo et al., 2020; Mackelprang et al., 2011; Monaco…

Continue Reading Comparative analysis of shotgun metagenomics and 16S rDNA sequencing of gut microbiota in migratory seagulls [PeerJ]

Single-cell transcriptomics reveals the brain evolution of web-building spiders

Animals for single-cell sequencing Adult samples of the aerial web-building spider (Hylyphantes graminicola) were collected from Anci district, Langfang, Hebei, China (39° 31.90’ N, 116° 38.15’ E) between September and October 2020. Collected spiders used for brain dissection were housed individually in a glass tube (Φ12 mm × 80 mm) at temperature- and humidity-controlled condition (24–26 °C and 50–60%…

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CRISPR/Cas9 bioluminescence-based assay for monitoring CFTR trafficking to the plasma membrane

Introduction Cystic fibrosis (CF) is a lethal, autosomal recessive disorder caused by mutations in the CFTR gene that results in aberrant transport of chloride, bicarbonate, and sodium ions (Kerem et al, 1989; Riordan et al, 1989; Rommens et al, 1989; Tabcharani et al, 1991; Poulsen et al, 1994; Stutts et…

Continue Reading CRISPR/Cas9 bioluminescence-based assay for monitoring CFTR trafficking to the plasma membrane

Catalytically inactive long prokaryotic Argonaute systems employ distinct effectors to confer immunity via abortive infection

Bacterial strains and phages Escherichia coli DH5a was routinely grown in Lysogeny broth (LB) medium and used for plasmid cloning, while E. coli BL21 (DE3) was used for protein production and the in vivo assays. E. coli phages T5, T7 and Lambda-vir are gifts from Shi Chen lab (Wuhan University,…

Continue Reading Catalytically inactive long prokaryotic Argonaute systems employ distinct effectors to confer immunity via abortive infection

Ingenio Kit-Amaxa 100 rxn, 0.2 cm Cuvette/Droppers

Description High Efficiency Electroporation– Deliver to hard to transfect cell lines and primary cells Compatible with Most Conventional Electroporation Devices – Use your existing system including the Ingenio® EZporator®, Lonza-Amaxa®, Bio-Rad® or Harvard BTX® Save Money and Reduce Research Costs Without Sacrificing Performance – Ingenio® Electroporation Solution is available as a stand-alone solution or…

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Identification of CCZ1 as an essential lysosomal trafficking regulator in Marburg and Ebola virus infections

Cells and viruses Haploid mSCs AN3–12 are a feeder independent clonal derivative of HMSc2 isolated from mice oocyte and maintained at IMBA52. The AN3–12 library and knocked out cells used for the haploid screening was obtained from IMBA (Austria)14,52. AN3–12 cells were validated by STR analysis. Haploid mES cells were…

Continue Reading Identification of CCZ1 as an essential lysosomal trafficking regulator in Marburg and Ebola virus infections

Investigating the miniaturization of DNA library prep for Illumina sequencing

Recent advancements in next-generation sequencing have led to substantial increases in throughput, coupled with reduced expenses. Due to these improvements, the process of library preparation has become even more of a financial and time constraint to high throughput sequencing core facilities. Nonetheless, with the help of automation in the form…

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Structure of human TRPM8 channel

Reagents and chemicals pFastBac Dual His6 MBP N10 TEV LIC cloning vector (5 C) was purchased from Addgene (Addgene plasmid no. 30123; www.addgene.org/30123). All restriction enzymes, the pFastBac1 vector, and DH10Bac cells were purchased from Thermo Fisher Scientific. Sf9 cells were purchased from ATCC. Q5® High-Fidelity DNA Polymerase, the Monarch® DNA…

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How to interpret the discrepancy of assignment rate in featurecounts using forward and reverse strand protocols

How to interpret the discrepancy of assignment rate in featurecounts using forward and reverse strand protocols 0 My RNAseq analysis pipeline is as follows: fastqc (read quality is good, some overrepresentation of adaptor sequence) -> trimmomatic (trimmed adaptor sequence, qc report after trimming suggests the overrepresented adaptors are gone) ->…

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Solved Molecular Biology \& Genetics – MOB673 Independent

Transcribed image text: Molecular Biology \& Genetics – MOB673 Independent Laboratory Project Functional Cloning of Tetracycline Resistance Markers in E. coli using Native Promoters – Plasmid Engineering \& Selection Aim: Functional migration of Tetracycline Resistance Marker from a donor plasmid to a recipient plasmid utilizing expression cloning and constitutive, native…

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Inactive S. aureus Cas9 downregulates alpha-synuclein and reduces mtDNA damage and oxidative stress levels in human stem cell model of Parkinson’s disease

Cloning of CRISPR/sgRNA lentiviral constructs with fluorescent selection markers A tetracycline-inducible promoter (TRE3G) was used to control the expression of S. aureus dCas9 in a lentiviral vector. To facilitate selection of cells by FACS, pHR:TRE3G-SadCas9-2xKRAB-p2a-tdTomato (Addgene ID #209298) was subcloned from a pHR:TRE3G-SadCas9-2xKRAB-p2a-zeo (A gift from Professor Stanley Qi), where zeocin…

Continue Reading Inactive S. aureus Cas9 downregulates alpha-synuclein and reduces mtDNA damage and oxidative stress levels in human stem cell model of Parkinson’s disease

Genomic Characteristics of Carbapenem-Resistant Klebsiella pneumoniae

Introduction Klebsiella pneumoniae (K. pneumoniae), which belongs to the family Enterobacteriaceae, is a common nosocomial pathogen that causes a variety of human infections, including bloodstream infection, respiratory tract infection, urinary tract infection, and abdominal infection.1,2 Carbapenems are generally considered to be a last choice for the treatment of multi-drug resistant…

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DNA damage induced by CDK4 and CDK6 blockade triggers anti-tumor immune responses through cGAS-STING pathway

Patients’ tumor tissues and clinical data This study recruited 125 breast cancer patients for immunohistochemical (IHC) analysis and 10 breast cancer patients for RNA-seq analysis from the Affiliated Tumor Hospital of Nantong University. Tumor tissues collected from surgical patients were fixed with formalin and embedded with paraffin and then examined….

Continue Reading DNA damage induced by CDK4 and CDK6 blockade triggers anti-tumor immune responses through cGAS-STING pathway

Plants | Free Full-Text | Guidelines for Performing CRISPR/Cas9 Genome Editing for Gene Validation and Trait Improvement in Crops

2.2. Design of Guide RNAs The purpose of this step is to design and select sgRNAs that will create the desirable edits in the target areas of the genome with high efficiency. Currently, there are many online software tools (online software without the need to be installed) for the design…

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Targeted knockout of a conserved plant mitochondrial gene by genome editing

Plant material and growth conditions Nicotiana tabacum cultivar Petit Havana was used for all experiments. The TALEN design and the TALEN-expressing line Nt-JF1006-30 were described previously19. For plant growth under sterile conditions, surface-sterilized seeds were germinated on Murashige and Skoog (MS) medium52 consisting of premixed MS salts and modified vitamins…

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Rapid discovery of high-affinity antibodies via massively parallel sequencing, ribosome display and affinity screening

Construct design To transcribe and translate sequenced DNA clusters on an Illumina flow cell, our DNA constructs contained the following elements: a P5 adaptor, followed by a 28 nt unique barcode, a 27 nt unstructured spacer (5p UNS v2), a ribosome binding site, start codon, protein coding region, TolAk short linker, 2×…

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DNA Analysis Products | NEB

Whether you are performing your first cloning experiment or constructing multi-fragment DNA assemblies, NEB® has the solution for you. Our high quality reagents are available for every workflow, including popular DNA assembly methods such as NEBuilder® HiFi DNA Assembly and NEBridge® Golden Gate Assembly. We also offer solutions for automation, site-directed…

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The University of Nebraska Just Helped Unlock the Final Secrets of the Corn Genome, And It Could Be a Breakthrough for Yield

An aerial view of a corn field on the edge of Lincoln, Neb., is a view into the future. Researchers have done what seemed impossible a decade ago: completing the sequencing of the entire corn genome. “If you ask any farmer, I think they will tell you the three things…

Continue Reading The University of Nebraska Just Helped Unlock the Final Secrets of the Corn Genome, And It Could Be a Breakthrough for Yield

Can T7 DNA Polymerase be used in other NEBuffers, including rCutSmart?

Whether you are performing your first cloning experiment or constructing multi-fragment DNA assemblies, NEB® has the solution for you. Our high quality reagents are available for every workflow, including popular DNA assembly methods such as NEBuilder® HiFi DNA Assembly and NEBridge® Golden Gate Assembly. We also offer solutions for automation, site-directed…

Continue Reading Can T7 DNA Polymerase be used in other NEBuffers, including rCutSmart?

capTEs enables locus-specific dissection of transcriptional outputs from reference and nonreference transposable elements

Cell culture All cell lines were grown in 6 cm dishes at 37 °C in a 5% CO2 incubator. The K562, MDA-MB-231 and HCT 116 cell lines were cultured in high-glucose DMEM supplemented with 10% fetal bovine serum and 1% penicillin-streptomycin antibiotics (pen-strep). NCM460 cells were cultured in RPMI 1640 medium supplemented…

Continue Reading capTEs enables locus-specific dissection of transcriptional outputs from reference and nonreference transposable elements

Can ssDNA oligonucleotides be combined and assembled with dsDNA fragments?

Whether you are performing your first cloning experiment or constructing multi-fragment DNA assemblies, NEB® has the solution for you. Our high quality reagents are available for every workflow, including popular DNA assembly methods such as NEBuilder® HiFi DNA Assembly and NEBridge® Golden Gate Assembly. We also offer solutions for automation, site-directed…

Continue Reading Can ssDNA oligonucleotides be combined and assembled with dsDNA fragments?

Genome-wide probing of eukaryotic nascent RNA structure elucidates cotranscriptional folding and its antimutagenic effect

Yeast strain and growth conditions S. cerevisiae strain BY4741 was grown in YPD media at 30 °C. Saturated cultures were diluted into 300 ml YPD medium with an optical density at 660 nm (OD660) of approximately 0.1 and then grown at 30°C with shaking at 250 rpm until mid-log phase with an OD660 of…

Continue Reading Genome-wide probing of eukaryotic nascent RNA structure elucidates cotranscriptional folding and its antimutagenic effect

NEBNext Ultra II RNA LIbrary Prep

One or more of these products are covered by patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc. For more information, please email us at gbd@neb.com. The use of these products may require you to obtain additional third party intellectual property rights for certain applications. ILLUMINA®, TRUSEQ®…

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NEBNext DNA Sample Preparation Reagents

Canada’s Michael Smith Genome Sciences Centre Validates NEBNext® DNA Sample Preparation Reagents From New England Biolabs For Use In Next Generation Sequencing Ipswich, MA (August 4, 2010) – New England Biolabs is pleased to announce that Canada’s Michael Smith Genome Sciences Centre has selected New England Biolabs as one of…

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Viruses | Free Full-Text | Optimizing the Conditions for Whole-Genome Sequencing of Avian Reoviruses

1. Introduction Avian orthoreoviruses (avian reoviruses, ARVs) are a persistent challenge to poultry producers in the United States and globally. Infection with ARVs has been associated with a variety of symptoms and syndromes in commercial poultry, including tenosynovitis/viral arthritis, enteric symptoms such as watery diarrhea, respiratory symptoms, myocarditis, viral hepatitis,…

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DNA sequencing at the picogram level to investigate life on Mars and Earth

All the extractions, dilutions and library preparation for sequencing were carried out inside the clean room environment of ISO (International Organization for Standardization) level 5 (ECSS-Q-ST-70-55C standard)32. An ISO 5 Class clean room is a semi-closed ultra clean environment that utilises High Efficiently Particle Air (HEPA) filtration systems to maintain…

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sgRNA Synthesis Reagents and Kits

Products and content are covered by one or more patents, trademarks and/or copyrights owned or controlled by New England Biolabs, Inc (NEB). The use of trademark symbols does not necessarily indicate that the name is trademarked in the country where it is being read; it indicates where the content was originally…

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RNA-seq Products | NEB

Whether you are performing your first cloning experiment or constructing multi-fragment DNA assemblies, NEB® has the solution for you. Our high quality reagents are available for every workflow, including popular DNA assembly methods such as NEBuilder® HiFi DNA Assembly and NEBridge® Golden Gate Assembly. We also offer solutions for automation, site-directed…

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DNA Ligase Selection Chart | NEB

Whether you are performing your first cloning experiment or constructing multi-fragment DNA assemblies, NEB® has the solution for you. Our high quality reagents are available for every workflow, including popular DNA assembly methods such as NEBuilder® HiFi DNA Assembly and NEBridge® Golden Gate Assembly. We also offer solutions for automation, site-directed…

Continue Reading DNA Ligase Selection Chart | NEB

T7 DNA Polymerase (unmodified) | NEB

Unit Definition One unit is defined as the amount of enzyme that will incorporate10 nmoles of dNTP into acid insoluble material in 30 minutes at 37°C. Reaction Conditions 1X T7 DNA Polymerase Reaction Buffer Supplement with Recombinant Albumin, Molecular Biology Grade Incubate at 37°C 1X T7 DNA Polymerase Reaction Buffer 20…

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EnGen Lba Cas12a (Cpf1) | NEB

EnGen® Lba Cas12a from Lachnospiraceae bacterium ND2006 is a site-specific DNA endonuclease guided by a single 41-44 nucleotide guide RNA (gRNA) (1). Targeting requires a gRNA complementary to the target site as well as a 5’ TTTV protospacer adjacent motif (PAM) on the DNA strand opposite the target sequence. Cleavage…

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Inter-bacterial mutualism promoted by public goods in a system characterized by deterministic temperature variation

Location and sample collection Samples were taken from 10 composting piles (between May and August 2020) at a food waste (FW) composting facility (Zhejiang, China; 30°52′51.50″N; 119°46′12.87″E). A consistent pre-treatment method was used for each pile. Specifically, about 8 tons of FW was collected every 3 days and subsequently crushed into…

Continue Reading Inter-bacterial mutualism promoted by public goods in a system characterized by deterministic temperature variation

Symmetric inheritance of parental histones governs epigenome maintenance and embryonic stem cell identity

The research in this study was conducted under the ethical approval of the Danish Regulatory Authority under project license 2018-15-0201-01520. Cell culture and differentiation assays WT, MCM2-2A, MCM2-R and POLE4-KO mouse ESCs used in this study were derived from the male, E14JU cell line with a 129/Ola background81. Genome editing…

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Staff Scientist, Molecular Enzymology Division

Currently, we are recruiting a Postdoc who will explore new areas in DNA polymerase enzymology by developing novel analytical, genetic and next generation sequencing techniques to understand new classes of DNA polymerases.  The New England Biolabs Postdoc Program trains emerging scientists to become independent investigators and leaders in their fields….

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Generation of inactivated IL2RG and RAG1 monkeys with severe combined immunodeficiency using base editing

Animals The gene targeting experiment was conducted on cynomolgus monkeys, which served as the experimental subjects. These monkeys were housed at Guangdong LANDAU Biotechnology Co. Ltd and TOPGENE Biotechnology Co. Ltd, where they received unrestricted access to water and were provided with a standard diet in accordance with standard care…

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Single-cell massively-parallel multiplexed microbial sequencing (M3-seq) identifies rare bacterial populations and profiles phage infection

Bacterial strains and growth conditions for eBW1 B. subtilis 168 and E. coli (MG1655) were streaked out from a frozen glycerol stock onto an LB plate and grown overnight at 37 °C. Following a night of growth, a single colony was picked and inoculated into 5 ml of LB broth and grown…

Continue Reading Single-cell massively-parallel multiplexed microbial sequencing (M3-seq) identifies rare bacterial populations and profiles phage infection

Selective enrichment of plasma cell-free messenger RNA in cancer-associated extracellular vesicles

Clinical samples and plasma preparation Blood samples from control individuals and patients with multiple myeloma, liver cancer, and lung cancer were obtained from Oregon Health and Science University (OHSU) by Knight Cancer Institute Biolibrary and Oregon Clinical and Translational Research Institute (OCTRI). All samples were collected under OHSU institutional review…

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MenT nucleotidyltransferase toxins extend tRNA acceptor stems and can be inhibited by asymmetrical antitoxin binding

MenAT1 sequence analysis Analysis of gene neighbourhoods for rv0078B (menA1) was performed using default settings in FlaGs (www.webflags.se/). Output sequences for MenA1 and cognate MenT1 homologues were then used to perform sequence alignments using MUSCLE (www.ebi.ac.uk/Tools/msa/muscle/), then formatted in Jalview (www.jalview.org/), sorting by pairwise alignment. Residues of interest were then…

Continue Reading MenT nucleotidyltransferase toxins extend tRNA acceptor stems and can be inhibited by asymmetrical antitoxin binding

CRISPR/Cas12a combined with RPA for detection of T. gondii in mouse whole blood | Parasites & Vectors

Mice and parasites The strains RH, Pru, wh3, and wh6, utilized in the present investigation, were generously provided by the Anhui Key Laboratory of Zoonoses, Anhui Medical University. The DNA of C. parvum and Babesia were procured from Shanghai Veterinary Research Institute, Chinese Academy of Agricultural Sciences. Furthermore, Plasmodium was…

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Rapid gene content turnover on the germline-restricted chromosome in songbirds

This work was carried out in accordance with ethical animal research requirements of Poland according to Polish law (the Act On the Protection of Animals used for Scientific or Educational Purposes, 15.01.2015, item 266, implementing Directive 2010/63/EU of the European Parliament and of the European Council of 22.09.2010). Experiments on…

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cp2k 2023.2 – Download, Browsing & More

cp2k 2023.2 – Download, Browsing & More | Fossies Archive “Fossies” – the Fresh Open Source Software Archive Contents of cp2k-2023.2.tar.bz2 (27 Jul 10:02, 68243435 Bytes) About: CP2K is a quantum chemistry and solid state physics software package that can perform atomistic simulations of solid state, liquid, molecular, periodic, material,…

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Molecular features driving condensate formation and gene expression by the BRD4-NUT fusion oncoprotein are overlapping but distinct

Cell culture HCC2429 cells were a kind gift from the Hamon Center for Therapeutic Oncology Research at UT Southwestern Medical Center. The cells were grown in RPMI 1640 medium (Thermo Fisher Scientific, #11875119) with addition of 10% heat-inactivated Fetal Bovine Serum (Thermo Fisher Scientific, #10-438-026) and 1% penicillin—streptomycin (Thermo Fisher…

Continue Reading Molecular features driving condensate formation and gene expression by the BRD4-NUT fusion oncoprotein are overlapping but distinct

Effects of Candidatus Liberibacter asiaticus infection on metagenome of Diaphorina citri gut endosymbiont

Psyllid sampling, tissue collection and DNA isolation Adult D. citri newly emerged (five days old) was initially harvested from Citrus Tachibana uninfected CLas in 2020 at Guangxi province, China, and was successively reared on Murraya paniculata at Guangxi Special Crops Research Institute for more than 15 generations. All cages and…

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Optimized minimal genome-wide human sgRNA library

sgRNA collection and enzyme cut site annotation All sgRNAs corresponding to potential NGG-containing target sites on the human (GRCh38/hg38) genome were calculated by a customized Perl script. The enzyme cut site was set at the 17 position from 5’ to 3’ of each sgRNA sequence. These sgRNAs have more than…

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Zambon Presents Results from the Two Phase 3 PROMIS Studies of CMS I-neb in Patients with Non-Cystic Fibrosis Bronchiectasis at World Bronchiectasis Conference 2023

Data show inhalation via the I-neb Adaptive Aerosol Delivery System of colistimethate sodium (CMS) results in reduction of pulmonary exacerbations in non-cystic fibrosis bronchiectasis (NCFB) patients compared to placebo The totality of the evidence from the Phase 3 PROMIS program supports a clinically meaningful benefit of CMS in reducing exacerbation…

Continue Reading Zambon Presents Results from the Two Phase 3 PROMIS Studies of CMS I-neb in Patients with Non-Cystic Fibrosis Bronchiectasis at World Bronchiectasis Conference 2023

Amphioxus adenosine-to-inosine tRNA-editing enzyme that can perform C-to-U and A-to-I deamination of DNA

Animals and cells Adult amphioxus B. japonicum collected from the sandy bottom of the sea near Qingdao, China, were cultured in aerated seawater at room temperature and fed with single-celled algae once a day. Zebrafish (D. rerio) aged 3 months and adult Japanese scallops (M. yessoensis) were purchased from Nanshan…

Continue Reading Amphioxus adenosine-to-inosine tRNA-editing enzyme that can perform C-to-U and A-to-I deamination of DNA

ResR/McdR-regulated protein translation machinery contributes to drug resilience in Mycobacterium tuberculosis

Bacterial strains and culture conditions Escherichia coli strain DH5α (Thermo Fisher) was used for the propagation of plasmids, whereas E. coli BL21 DE3 (Novagen) was used for the expression and purification of ResR/McdR protein. Mtb Erdman was obtained from Dr. Ramandeep Singh at THSTI, India, and Mtb H37Rv mc2 790242…

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The Report provides complete market analysis of Biotechnology market and reports related to this market, forecasted for period from 2023

PRESS RELEASE Published July 10, 2023 Biotechnology Market The  Biotechnology market was valued at 528940 million in 2023 and is projected to reach US$ 856060 million by 2030, at a CAGR of 7.1% during the forecast for period from 2023 – 2030.  The biotechnology market research report provides insights into…

Continue Reading The Report provides complete market analysis of Biotechnology market and reports related to this market, forecasted for period from 2023

Single-cell quantification of ribosome occupancy in early mouse development

PDMS chip fabrication Moulds were 3D-printed by Proto Labs with WaterShed XC 11122 at high resolution (Supplementary Fig. 1). Reusable moulds were assembled by taping 3D-printed molds to glass slides (5″ × 4″; Ted Pella). Sylgard 184 PDMS monomer and curing agent (4019862, Ellsworth Adhesives) were mixed at a 10:1…

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Regulation of Molecular Genetic Engineering Must Be Evidence-Based

“Genetic Engineering” (GE) has been practiced by humans for more than 10,000 years, first by selecting and hybridizing plants. For almost half a century, newer molecular GE techniques have been used to modify plants, animals, bacteria and other organisms. Scientists can move desired genes virtually at will from one organism…

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Whole genome analysis for 163 gRNAs in Cas9-edited mice reveals minimal off-target activity

Allele design and guide selection For multi-exon genes, a critical region (one or more exons) was identified as shared among all annotated full-length transcripts whose removal was predicted to result in a frame-shift mutation and introduction of premature stop codon greater than 50-nt from the final splice junction increasing the…

Continue Reading Whole genome analysis for 163 gRNAs in Cas9-edited mice reveals minimal off-target activity

Prokaryotic winged helix domains as dsDNA adenine methylation sensors

Abstract Winged helix (wH) domains, also termed winged helix-turn-helix (wHTH) domains, are widespread in all kingdoms of life, and have diverse roles. In the context of DNA binding and DNA modification sensing, some eukaryotic wH domains are known as sensors of non-methylated CpG. In contrast, the prokaryotic wH domains in…

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acCRISPR: an activity-correction method for improving the accuracy of CRISPR screens

acCRISPR framework acCRISPR performs essential gene identification by calculating two scores for each sgRNA, namely the cutting score (CS) and the fitness score (FS). CS and FS are the log2-fold change of sgRNA abundance in the appropriate treatment sample with respect to that in the corresponding control sample (see Supplementary…

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switching from illumination to NEB prep kits

switching from illumination to NEB prep kits 0 Hi all, we already have built some pipelines for germline / somatic calling on WES data ( the library prep kits used were from Illumina). now we want to switch to using NEB library kits, and I would like to know (rom…

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Restriction Digestion Planning for RFLP Q5. What are

Transcribed image text: Restriction Digestion Planning for RFLP Q5. What are restriction enzymes and recognition sites? What is their function in nature? How can a mutation alter a recognition site? [10pts] Q6. Appendix I shows the restriction sites for NEB enzymes for the complete PTC gene. Choose a restriction enzyme…

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Phylogenomic analysis supports Mycobacterium tuberculosis transmission between humans and elephants

1. Introduction Tuberculosis (TB) is a significant global burden and is widely reported to be a major public health and economic problem, costing the world $617 billion between 2000 and 2015 and projected to cost $1 trillion between 2015 and 2030 (1). It is the second leading cause of death…

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Histone modifications regulate pioneer transcription factor cooperativity

Protein expression, mutagenesis and purification Xenopus laevis histones for nucleosome assembly were overexpressed in the Escherichia coli BL21(DE3) pLysS strain and purified from inclusion body as previously described48. The cells were grown in LB medium at 37 °C and induced with 1 mM IPTG when OD600 reached 0.6. After 3 h of expression,…

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bbduk command line options for EM-seq (NEB)

Hi I would like to use bbduk to replicate the command line flags used by trim_galore for EM-Seq (NEB). The bismark user guide recommends this for trim_galore –clip_R1 10 –clip_R2 10 –three_prime_clip_R1 10 –three_prime_clip_R2 10 Explanation of the trim_galore parameters –clip_R1 <int> Instructs Trim Galore to remove <int> bp from…

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Activation of the urotensin-II receptor by remdesivir induces cardiomyocyte dysfunction

Study design The overall goal of this study was to explore the molecular mechanisms of remdesivir-related cardiotoxicity in anti-COVID-19 therapy. We first performed the GPCR screening using major anti-COVID-19 drugs as ligands (n = 3 per GPCR). We validated the results of GPCR screening by concentration-response analysis and determined the EC50 and…

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In-depth Temporal Transcriptome Profiling of Monkeypox and Host Cells using Nanopore Sequencing

Figure 1 shows the detailed workflow of the study. Fig. 1 General overview of the study. Briefly, MPXV was isolated from a skin lesion and then was used to infect CV-1 cells. After the designated infection times, total RNA was isolated and sequenced using direct cDNA sequencing protocol on ONT’s MinION platform….

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Amplification Biases and Consistent Recovery of Loci in a Double-Digest RAD-seq Protocol | PLOS ONE – Molecular Biology Protocol – Restriction Digest of Plasmid …

Abstract A waxing variety of “genotype-by-sequencing” (GBS) methods use restriction enzymes and high throughput DNA ordering to generation data for a set of genomic loci, allowing the simultaneous discovery press genotyping of thousands of polymorphisms in a set of multiplexed samples. We evaluated a “double-digest” restriction-site associated DNA sequencing (ddRAD-seq)…

Continue Reading Amplification Biases and Consistent Recovery of Loci in a Double-Digest RAD-seq Protocol | PLOS ONE – Molecular Biology Protocol – Restriction Digest of Plasmid …

Roles of adenine methylation in the physiology of Lacticaseibacillus paracasei

Bacterial strains and cultivation Twenty-eight L. paracasei isolates (including L. paracasei Zhang) and a pglX gene-inactivated strain of L. paracasei Zhang were obtained from the Key Laboratory of Dairy Biotechnology and Engineering, Ministry of Education, at the Inner Mongolia Agricultural University of China. For strain activation, the bacteria were cultivated…

Continue Reading Roles of adenine methylation in the physiology of Lacticaseibacillus paracasei

Optimized metrics for orthogonal combinatorial CRISPR screens

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…

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Hybrids of RNA viruses and viroid-like elements replicate in fungi

Ribozyme search of the Sequence Read Archive Observing that ribozymes are sufficiently short to be captured on a short sequence read (less than 100 nt), we reasoned it will be possible to screen large volumes of sequencing data to identify libraries potentially containing ribozyme agents. To this end we adapted…

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Highly-multiplexed and efficient long-amplicon PacBio and Nanopore sequencing of hundreds of full mitochondrial genomes | BMC Genomics

Laboratory methods Tissue samples of lizards were collected over the course of several field seasons in Indonesia under appropriate permits as part of a biotic survey of mountains across the island of Sulawesi. Liver tissues were dissected and stored in RNA-later, which were incubated at ambient temperature for 24 to…

Continue Reading Highly-multiplexed and efficient long-amplicon PacBio and Nanopore sequencing of hundreds of full mitochondrial genomes | BMC Genomics